Kang Zhang Laboratory Publications

Publications

Original Articles

1. Kaplan, G., Zhang, K. and Beverley, S. (1987). Sequence and S1 mapping of the 5' end of the DHFR-TS gene of Leishmania major. Nucleic Acid Research 15, 3369.
   
   Abstract: The 5' structure of mRNA transcribed from the dihydrofolate reductase- thymidylate synthase (DHFR-TS) gene of the protozoan parasite Leishmania major has been characterized. S1 nuclease mapping identifies a heterogenous 5' structure which is not affected by growth phase or developmental stage. The DNA sequence of the 5' region of the DHFR-TS gene does not reveal homology with other trypanosomatid genes, eukaryotic consensus genetic elements, or the mammalian DHFR promoter element. This latter finding is especially significant as we show that the 5' region of the E. coli DHFR gene exhibits homology to the mammalian DHFR promoter element, despite their greater evolutionary distance. (full text pdf)
   
   
2. Kaplan, G., Zhang, K. and Beverley, S. (1990). Nuclease mapping and DNA sequence analysis of transcripts from the di-hydrofolate reductase thymidylate synthase (R) region of Leishmania major. Nucleic Acid Research 18, 6399.
   
   Abstract: Trypanosomatid protozoan parasites utilize a number of nonstandard mechanisms in expressing their genes. To probe these phenomena in a genetically accessible system, we have mapped termini of eight transcripts arising from the amplified R region including the DHFR-TS gene of methotrexate-resistant Leishmania major. Poly(A)+ RNAs transcribed from the DHFR-TS-coding strand exhibit features similar to those observed around other trypanosomatid protein-coding genes. These include close spacing, the presence of a transpliced miniexon on the 5' termini, heterogeneity at both 5' and 3' ends, and in some cases S1 nuclease protection of intertranscript regions. Other than the splice acceptor site, no consensus sequence elements associated with either 5' or 3' ends were detected, although polydinucleotide tracts tended to be near inter-transcript regions. Two poly(A)+ RNAs transcribed from the opposite strand of the upstream flanking regions lacked the miniexon. Sequencing of DNA encoding the overlapping 1.7 kb opposite strand transcripts (one bearing and one lacking the miniexon, both found on polysomes) revealed no reading frames likely to encode proteins, suggesting that at least some of these RNAs could be nonfunctional by-products of RNA processing. (full text pdf)
   
   
3. Perkins, L.A., Doctor, J., Zhang, K., Stinson, L., Perrimon, N., Craig, E. (1990). Molecular and developmental characterization of the heat shock cognate 4 gene of Drosophila Melanogaster. Mol. Cell. Biol. 10, 3232.
   

   Abstract: The Drosophila heat shock cognate gene 4 (hsc4), a member of the hsp70 gene family, encodes an abundant protein, hsc70, that is more similar to the constitutively expressed human protein than the Drosophila heat-inducible hsp70. Developmental expression revealed that hsc4 transcripts are enriched in cells active in endocytosis and those undergoing rapid growth and changes in shape. (full text pdf)

4. Zhang, K., Chaillet, R., Perkins, L.A., Halazonetis, T. and Perrimon, N. (1990). The Drosophila homolog of the mammalian jun oncogene is expressed during embryonic development and activates transcription in mammalian cells. Proc. Natl. Acad. Sci. USA. 87, 6281.
   
   Abstract: By means of low-stringency cross-species hybridization to Southern DNA blots, human c-jun sequences were used to identify a unique Drosophila melanogaster locus (Djun). The predicted DJun protein is highly homolog ous to members of the mammalian Jun family in both the DNA binding and leucine zipper regions. Djun was mapped by in situ hybridization to position 46E of the second chromosome. It encodes a 1.7-kilobase transcript constitutively expressed at all developmental stages. Functionally, Djun in cooperation with mouse c-fos can trans-activate activator protein 1 DNA binding site when introduced into mammalian cells. Taken together, these data suggest that Djun, much like its mammalian homolog , may activate transcription of genes involved in regulation of cell growth, differentiation, and development. Furthermore, the identification of Djun allows one to exploit the genetics of Drosophila to identify genes in signal transduction pathways involving Djun and thus c-jun. (full text pdf)
   
   
5. Zhang, K., Smouse, D. and Perrimon, N. (1991). The crooked neck gene of Drosophila contains a motif found in a family of yeast cell cycle genes. Genes & Development 5, 1080.
   
   Abstract: The crooked neck (crn) gene of Drosophila encodes a protein of 702 amino acids and contains 16 tandemly arranged copies of a 34-amino-acid repeat that is similar to the tetratrico peptide repeat (TPR). Multiple copies of the TPR motif have also been found in a family of yeast genes, including several members that are necessary for cell division. TPR-containing proteins encoded by the yeast genes CDC16, CDC23, and nuc2+ are required for progression through the G2/M transition of the cell cycle. Loss of zygotic expression of crn causes defects in the proliferation of brain neuroblasts and results in the absence of identified neuronal lineages in the central and peripheral nervous systems. The sequence similarity and mutant phenotypes are consistent with a cell cycle requirement for the crn gene product.
   
   
6. Rutledge, B.*, Zhang, K.*, Bier, E., Jan, Y. and Perrimon, N. (1992) The Drosophila spitz gene encodes a putative EGF-like growth factor involved in dorsal-ventral axis formation and neurogenesis. Genes & Dev. 6, 1503. * co-first authors
   
   Abstract: We describe the molecular characterization of the Drosophila gene spitz (spi), which encodes a putative 26-kD, EGF-like transmembrane protein that is structurally similar to TGF-alpha. Temporal and spatial expression patterns of spi transcripts indicate that spi is expressed throughout the embryo. Examination of mutant embryos reveals that spi is involved in a number of unrelated developmental choices, for example, dorsal-ventral axis formation, glial migration, sensory organ determination, and muscle development. We propose that spi may act as a ligand for cell-specific receptors, possibly rhomboid and/or the Drosophila EGF receptor homolog.
   
   
7. Zhang, K., Bither, P.P., Park, R., Donoso, L.A., Seidman, J.G., and Seidman, C.E. (1994). A dominant Stargardt's macular dystrophy locus maps to chromosome 13q34. Archives of Ophthalmology 112, 759.
   
   Abstract: OBJECTIVE: To identify the chromosomal location of a mutated gene that causes an autosomal dominant Stargardt's macular dystrophy. METHODS: Ocular examinations were performed on 67 members of a large kindred to identify those with macular dystrophy. DNA analyses defined the genotype of all family members at 49 polymorphic loci. Linkage between the gene defect responsible for this macular dystrophy and each polymorphic locus was assessed by lodscore calculations. RESULTS: Diminished visual acuity and funduscopic abnormalities were found in 29 family members, which was diagnostic of macular dystrophy. Genetic analyses demonstrated that polymorphic loci from chromosome 13 band q34 were linked to the gene defect in this family. Haplotype analyses localized the disease locus to an 8-centimorgan interval between loci D13S159 and D13S158/D13S174. CONCLUSION: A disease locus responsible for an autosomal dominant Stargardt's macular dystrophy is located on chromosome 13 band q34. Identification of the mutated gene at this locus will lead to a better understanding of macular degeneration.
   
   
8. Yasuaki, H., Duh, E., Zhang, K., Robinson, G. S., Aiello, L.P. (1998). Transcription factors Sp1 and Sp3 alter vascular endothelial growth factor receptor expression through a novel recognition sequence. J. Biol. Chem. 273, 19294.
   
   Abstract: Kinase domain receptor (KDR) is a high affinity, endothelial cell-specific, autophosphorylating tyrosine kinase receptor for vascular endothelial growth factor. This transcriptionally regulated receptor is a critical mediator of endothelial cell (EC) growth and vascular development. In this study, we identify a DNA element modulating KDR promoter activity and evaluate the nuclear binding proteins accounting for a portion of the cell-type specificity of the region. KDR promoter luciferase activity was retained within -85/+296 and was 10-30-fold higher in EC than non-EC. Electrophoretic mobility shift assays demonstrated specific nuclear protein binding to -85/-64, and single point mutations suggested important binding nucleotides between -79/-68 with five critical bases between -74/-70 (5'-CTCCT-3'). DNA-protein complexes were displaced by Sp1 consensus sequence oligodeoxynucleotides and supershifted by Sp1- and Sp3-specific antibodies. Sp1 and Sp3 protein in EC nuclear extracts bound the -79/-68 region even when all surrounding classic Sp1 recognition sites were removed. Sp1 protein in nuclear extracts was 4-24-fold higher in EC than non-EC, whereas Sp3 was 3-7-fold higher. Sp1/Sp3 ratios in EC were 2-10-fold higher. Overexpression of Sp1 protein increased KDR promoter activity 3-fold in both EC and non-EC, whereas simultaneous co-expression of Sp3 attenuated this response. An Sp1 consensus sequence cis element "decoy" reduced EC KDR promoter activity and mRNA expression by 85 and 69%, respectively. An antisense phosphorothioate oligodeoxynucleotide to Sp1 inhibited Sp1 and KDR protein expression by 66 and 68%, respectively, without changing Sp3 protein expression. These data illustrate that Sp1 and Sp3 modulate KDR promoter activity through a novel recognition binding sequence. However, since Sp1-mediated promoter activation is attenuated by Sp3, endothelial selective KDR promoter activity may be partially regulated by variations in the Sp1/Sp3 ratio. (full text pdf)
   
   
9. Zhang, K., and Jampel, H. (1999). Discordance of congenital glaucoma in monozygous twins. American Journal of Ophthalmology 128, 97.
   
   Abstract: PURPOSE: To report a case of primary infantile glaucoma occurring in only one of two identical twins. METHODS: Case reports. RESULTS: Clinical features of bilateral primary infantile glaucoma in one of the two twins at age 16 months are described. Polymorphic DNA markers were used to establish identical twinship. CONCLUSION: Our finding suggests that nongenetic factors must be considered in determining the origin of primary infantile glaucoma.
   
   
10. Kniazeva, M., Chiang, M.F., Cutting, G., Zack, D. J., Han, M., and Zhang, K. (1999). Clinical and Genetic studies of a cone-rod dystrophy with features of Stargardt'disease. Ophthalmic Genetics 20, 71.
    
    Abstract: Cone-rod dystrophy (CORD) and Stargardt disease (STGD) are two hereditary retinal dystrophies with similarities to age-related macular degeneration. Cone-rod dystrophies are a group of degenerative disorders resulting in decreased visual acuity and color vision, attenuated electroretinographic (ERG) responses, and atrophic macular lesions. Autosomal dominant, autosomal recessive, and X-linked forms of cone-rod dystrophy have been reported. Stargardt disease is characterized by reduced visual acuity, atrophic macular changes, prominent 'flavimaculatus flecks' in the pigment epithelium of the posterior retina, and a virtually pathognomic 'dark choroid' pattern on fluorescein angiography. Stargardt disease is classically inherited as an autosomal recessive trait, although numerous families have been described in which features of Stargardt disease are transmitted in an autosomal dominant manner. We have identified a new kindred with autosomal dominant cone-rod dystrophy with features of Stargardt-like disease. Detailed clinical evaluation, genotype analysis, and linkage analysis were performed. Fluorescein angiography revealed a 'dark choroid' pattern in three affected subjects. Electroretinography disclosed markedly reduced scotopic and photopic responses in three affected individuals. Genetic analysis revealed linkage to known loci for cone-rod dystrophy (CORD7) and Stargardt-like disease (STGD3) on chromosome 6q14. A peak lod score of 3.3 was obtained with the marker D6S280 at straight theta =0.010. A physical map was constructed by screening a YAC library with short tandem repeat markers in the region. Screening of a candidate gene, the rho1 subunit of the GABA receptor, failed to reveal any mutations.
    
    
11. Kniazeva, M., Chiang, M.F., Morgan, B., Anduze, A.L., Zack, D.J., Han, M., and Zhang, K. (1999). A new locus for autosomal dominant Stargardt-like disease maps to chromosome 4. American Journal Human Genetics. 64, 1394.
    
    Abstract: Stargardt disease (STGD) is the most common hereditary macular dystrophy and is characterized by decreased central vision, atrophy of the macula and underlying retinal-pigment epithelium, and frequent presence of prominent flecks in the posterior pole of the retina. STGD is most commonly inherited as an autosomal recessive trait, but many families have been described in which features of the disease are transmitted in an autosomal dominant manner. A recessive locus has been identified on chromosome 1p (STGD1), and dominant loci have been mapped to both chromosome 13q (STGD2) and chromosome 6q (STGD3). In this study, we describe a kindred with an autosomal dominant Stargardt-like phenotype. A genomewide search demonstrated linkage to a locus on chromosome 4p, with a maximum LOD score of 5.12 at a recombination fraction of.00, for marker D4S403. Analysis of extended haplotypes localized the disease gene to an approximately 12-cM interval between loci D4S1582 and D4S2397. Therefore, this kindred establishes a new dominant Stargardt-like locus, STGD4. (full text pdf)
    
    
12. Zhang, K., Garibaldi, D., Kniazeva, M., Chiang, M., Sunness, J., Dean, M., Allikmets, R. (1999) A novel mutation of the ABCR gene in four patients with recessive Stargardt macular dystrophy. America. Journal of Ophthalmology 128, 720.
    
    Abstract: PURPOSE: To identify additional mutations in the ABCR gene and describe the clinical features of four affected siblings with autosomal recessive Stargardt disease. METHODS: A cohort of eight siblings was identified for study. Four of these individuals were diagnosed with Stargardt disease based on clinical evaluation and fluorescein angiography. Blood samples were obtained from seven of eight siblings, including all those affected. All 50 exons of the ABCR gene were analyzed by single-stranded confirmation polymorphism analysis, followed by direct sequencing of observed variants, to identify mutations in the ABCR gene. RESULTS: We identified a previously unreported kindred of eight siblings, four of whom had mutations in both of their ABCR alleles. A previously described G-to-C transversion of nucleotide 2588, predicting a Gly863Ala amino acid substitution, and a novel G-to-A transition of nucleotide 161, resulting in a Cys54Tyr substitution, were identified. These mutations co-segregated with the affected members of this family. Three of the siblings demonstrated clinical features characteristic of classic Stargardt disease, with bilateral regions of macular atrophy associated with yellow-white "flavimaculatus" flecks in the posterior pole at the level of the retinal pigment epithelium. The fourth affected sibling showed features of early Stargardt disease, with a beaten-bronze appearance to both maculas, as well as perimacular flecks. In all four affected patients, fluorescein angiography showed a characteristic peripheral dark choroid. CONCLUSIONS: We have identified both a previously described and a novel mutation in the ABCR gene in four patients with autosomal recessive Stargardt disease. In-depth knowledge of the ABCR mutation spectrum in patients with Stargardt disease will provide for more efficient screening and may provide potential therapies for Stargardt disease and other retinal diseases.
    
    
13. Zhang, K., Kniazeva, M., Han, M., Dean, M., Allikmets, R. (1999). The ABCR gene in dominant and recessive Stargardt's disease: a genetic pathway in macular degeneration. Genomics 60, 234.
    
    Abstract: Stargardt disease (STGD) is a juvenile-onset macular dystrophy and can be inherited in an autosomal recessive or in an autosomal dominant manner. Genes involved in dominant STDG have been mapped to human chromosomes 13q (STGD2) and 6q (STGD3). Here, we identify a new kindred with dominant STGD and demonstrate genetic linkage to the STGD3 locus. Because of a more severe macular degeneration phenotype of one of the patients in this family, the gene responsible for the recessive STGD1, ABCR, was analyzed for sequence variants in all family members. One allele of the ABCR gene was shown to carry a stop codon-generating mutation (R152X) in three family members, including the one patient who had inherited also the dominant gene. A grandparent of that patient with the same ABCR mutation developed age-related macular degeneration (AMD), consistent with our earlier observation that some variants in the ABCR gene may increase susceptibility to AMD in the heterozygous state. Based on these results, we propose that there is a common genetic pathway in macular degeneration that includes genes for both recessive and dominant STGD.
    
    
14. Kniazeva, M., Traboulsi, E.L., Stefko, S.T., Gorin, M.B., Blaschak, C.L.,Cutting,G. Han, M., and Zhang, K.. (2000). A new locus for dominant drusen with macular degeneration maps to chromosome 6q. American Journal of Ophthalmology 130, 197.
    
    Abstract: PURPOSE: To report the localization of a gene causing drusen and macular degeneration in a previously undescribed North American family. METHODS: Genetic mapping studies were performed using linkage analysis in a single family with drusen and atrophic macular degeneration. RESULTS: The clinical manifestations in this family ranged from fine macular drusen in asymptomatic middle-aged individuals to atrophic macular lesions in two children and two elderly patients. We mapped the gene to chromosome 6q14 between markers D6S2258 and D6S1644. CONCLUSIONS: In a family with autosomal dominant drusen and atrophic macular degeneration, the gene maps to a 3.2-cM region on chromosome 6q14. This locus appears to be distinct from, but adjacent to, the loci for cone-rod dystrophy 7 (CORD7) and North Carolina macular dystrophy (MCDR1). Future identification of the gene responsible for the disease in this family will provide a better understanding of macular degeneration.
    
    
15. Stefko, S. T., Zhang, K., Gorin, M.B., Elias I. Traboulsi, E.I. (2000). Clinical Spectrum of Chromosome 6 Linked Autosomal Dominant Drusen and Macular Degeneration. Am. J. Ophthalmology130, 203-8.
    
    Abstract: PURPOSE: To describe the clinical phenotype and the intrafamilial variation in retinal findings in a North American family with an autosomal dominant drusen disorder that maps to chromosome 6q14. METHODS: Ophthalmic examinations were carried out on participating family members. Fundus photographs were obtained whenever possible. Electroretinography was performed on the proband and her father. Blood was drawn for DNA analysis. RESULTS: Twelve family members had drusen and/or atrophic macular degeneration. The disease in asymptomatic young adults is characterized by fine drusen that are most conspicuous in the macula. The proband presented at 3 years of age with atrophic maculopathy and drusen. Her cousin was found to have atrophic macular lesions and drusen in the first year of life. Two older affected individuals have reduced vision from cicatricial and atrophic macular changes. The gene for the disease was mapped to chromosome 6q14 and appears to be adjacent to but distinct from the locus for North Carolina macular dystrophy. CONCLUSIONS: There is extreme variability in the clinical expression of this dominant form of drusen and macular degeneration. Most young adults have fine macular drusen and good vision. Affected infants and children may have congenital atrophic maculopathy and drusen. There is historical evidence of progression of the disease in late adulthood with moderate visual loss.
    
    
16. Allikmets,R#., Lewis, R.A.#, Steiner, K.#, Dean, M.#, Bernstein, P#, Seddon, J.M.#;. Zhang, K.#, Udar, N.S.#, Simonelli, F.#, Ingvast, S.#, Souied, E.#, Maugeri, A.#, Paloma, E.#, Kermani, S.#, and International ABCR screening Consortium. (2000). Further evidence for an association of the ABCR alleles with age-related macular degeneration. Am J Human Genetics 67, 487. #co-first authors
    
    Abstract: Age-related macular degeneration (AMD) accounts for >50% of the registered visual disability among North American and Western European populations and has been associated both with environmental factors, such as smoking, and with genetic factors. Previously we have reported disease-associated variants in the ABCR (also called ABCA4) gene in a subset of patients affected with this complex disorder. We have now tested our original hypothesis, that ABCR is a dominant susceptibility locus for AMD, by screening 1,218 unrelated AMD patients of North American and Western European origin and 1,258 comparison individuals from 15 centers in North America and Europe for the two most frequent AMD-associated variants found in ABCR. These two sequence changes, G1961E and D2177N, were found in one allele of ABCR in 40 patients ( approximately 3.4%), and in 13 control subjects ( approximately 0.95%). Fisher's two-sided exact test confirmed that these two variants are associated with AMD at a statistically significant level (P<.0001). The risk of AMD is elevated approximately threefold in D2177N carriers and approximately fivefold in G1961E carriers. The identification of a gene that confers risk of AMD is an important step in unraveling this complex disorder. (full text pdf)
    
    
17. Zhang, K., Kniazeva, M., Han, M., Li, W., Yu, Z., Yang, Z., Li, Y., Metzker, M. L., Allikmets, R. L., Zack, D. J., Kakuk, L. E., Lagali, P. S., Wong, P. W., MacDonald, I. M., Sieving, P. A., Figueroa, D., Austin, C. P., Robert J. Gould, R. J., Ayyagari, R., Petrukhin, K. (2001). A five base-pair deletion in the ELOVL4 gene is associated with two related forms of autosomal dominant macular dystrophy. Nature Genetics 27, 89.
    
    Abstract: Stargardt-like macular dystrophy (STGD3, MIM 600110) and autosomal dominant macular dystrophy (adMD) are inherited forms of macular degeneration characterized by decreased visual acuity, macular atrophy and extensive fundus flecks. Genetic mapping data suggest that mutations in a single gene may be responsible for both conditions, already known to bear clinical resemblance. Here we limit the minimum genetic region for STGD3 and adMD to a 0.6-cM interval by recombination breakpoint mapping and identify a single 5-bp deletion within the protein-coding region of a new retinal photoreceptor-specific gene, ELOVL4, in all affected members of STGD3 and adMD families. Bioinformatic analysis of ELOVL4 revealed that it has homology to a group of yeast proteins that function in the biosynthesis of very long chain fatty acids. Our results are therefore the first to implicate the biosynthesis of fatty acids in the pathogenesis of inherited macular degeneration.
    
    
18. Identification and functional consequences of a new mutation (E155G) in GCAP1 causing autosomal dominant cone dystrophy. Wilkie, S. E., Li, Y., Deery , E. C., Newbold, R., Garibaldi, D. C., Bateman, J. B., Zhang, H., Zack, D. J., Bhattacharya, S. S., Warren, M. J., Hunt, D. M., and Zhang, K. (2001). Am J Human Genetics 69, 471.
    
    Abstract: Mutations in the gene for guanylate cyclase-activating protein-1 (GCAP1) (GUCA1A) have been associated with autosomal dominant cone dystrophy (COD3). In the present study, a severe disease phenotype in a large white family was initially shown to map to chromosome 6p21.1, the location of GUCA1A. Subsequent single-stranded conformation polymorphism analysis and direct sequencing revealed an A464G transition, causing an E155G substitution within the EF4 domain of GCAP1. Modeling of the protein structure shows that the mutation eliminates a bidentate amino acid side chain essential for Ca2+ binding. This represents the first disease-associated mutation in GCAP1, or any neuron-specific calcium-binding protein within an EF-hand domain, that directly coordinates Ca2+. The functional consequences of this substitution were investigated in an in vitro assay of retinal guanylate cyclase activation. The mutant protein activates the cyclase at low Ca2+ concentrations but fails to inactivate at high Ca2+ concentrations. The overall effect of this would be the constitutive activation of guanylate cyclase in photoreceptors, even at the high Ca2+ concentrations of the dark-adapted state, which may explain the dominant disease phenotype. (full text pdf)
    
    
19. Li, Y., Marcos, I., Borrego, S., Yu, Z., Zhang, K., Antinolo, G. (2001). Evaluation of the ELOVL4 gene in families with Retinitis Pigmentosa linked to the RP25 locus. J. Med. Genetics 38, 478.
    
    Abstract: Retinitis pigmentosa (RP) is the most common form of retinal dystrophy. Patients present with night blindness and progressive narrowing of the visual field, eventually leading to central vision loss. Fundus examination usually shows bone spicula pigmentation, attenuation of blood vessels in the retina, and waxy pallor of the optic disc. Typically, the electroretinogram is notably diminished or even abolished. (full text pdf)
    
    
20. Zhang, K., Garibaldi, D., Li, Y., Green, W. R., Zack, D.J. (2002). Butterfly-shaped pattern dystrophy: A genetic, clinical and histopathologic report. Arch. Ophthalmology 120, 485.
    
    Abstract: OBJECTIVES: To identify the disease-causing mutation in a large family segregating dominantly inherited butterfly-shaped pattern dystrophy (BPD) and to describe the microscopic pathological changes observed in a member of this family. METHODS: Seventeen individuals at risk for dominantly inherited BPD in a family were examined and blood samples obtained. Linkage analysis and mutation screening of the human retinal degeneration slow (RDS)/peripherin locus were performed. Light and electron microscopic examinations were performed on 1 postmortem eye of 1 affected individual. RESULTS: Four individuals demonstrated macular degenerative changes with diminished visual acuity, and 3 others exhibited early signs of atrophy without visual deficits. Microscopic examination of the left eye of 1 patient revealed an area of total loss of the retinal pigment epithelium (RPE) and photoreceptor cell layer with intact choriocapillaris and lipofuscin-containing cells in the subretinal space. Outside the area of RPE atrophy, the RPE was greatly distended by lipofuscin. The disease locus in this family was mapped to 6p21.2, the region of the RDS/peripherin gene. Further analysis identified a G-->A change at nucleotide position 637 of RDS/peripherin, predicting a novel Cys213Tyr substitution in all affected members of the family. CONCLUSIONS: This study describes a new RDS/peripherin mutation for BPD and provides the first combined genetic-pathological study of this condition, to our knowledge. CLINICAL RELEVANCE: Accumulation of lipofuscin in RPE is a prominent feature of several retinal disorders, including age-related macular degeneration. Further elucidation of the cellular and molecular mechanism of BPD may provide insight into pathogenesis and lead to novel treatment approaches for this and other macular degenerations. (full text pdf)
    
    
21. Bernstein, P. S., Tammur, J., Singh, N., Hutchinson, A., Dixon, M., Pappas, C. M., Zabriskie, N. A., Zhang, K., Petrukhin, K., Leppert, M., Allikmets, R., (2001). Diverse macular dystrophy phenotypes caused by a novel complex mutation in the ELOVL4 gene. Investigative Ophthalmology and Visual Science 42, 3331.
    
    Abstract: PURPOSE: A 5-bp deletion in ELOVL4, a photoreceptor-specific gene, has been associated with autosomal dominant (ad) macular dystrophy phenotypes in five related families, in which phenotypes range from Stargardt-like macular dystrophy (STGD3; Mendelian Inheritance in Man 600110) to pattern dystrophy. This has been the only mutation identified in ELOVL4 to date, which is associated with macular dystrophy phenotypes. In the current study, the potential involvement was investigated of an ELOVL4 gene variation in adSTGD-like and other macular dystrophy phenotypes segregating in a large unrelated pedigree from Utah (K4175). METHODS: The entire open reading frame of the ELOVL4 gene was analyzed by direct sequencing in a proband from the K4175 family. The combination of denaturing high-performance liquid chromatography (DHPLC) analysis and direct sequencing of all available family members was used to further assess segregation of identified ELOVL4 variants in the pedigree. RESULTS: A complex mutation, two 1-bp deletions separated by four nucleotides, was detected in all affected members of the family. The mutation results in a frameshift and the truncation of the ELOVL4 protein, similar to the effect of the previously described 5-bp deletion. CONCLUSIONS: The discovery of a second mutation in the ELOVL4 gene segregating with macular dystrophy phenotypes confirms the role of this gene in a subset of dominant macular dystrophies with a wide range of clinical expressions and suggests a role for modifying genes and/or environmental factors in the disease process. (full text pdf)
    
    
22. Ayyagari, R., Zhang, K., Hutchinson, A, Yu, Z, Swaroop, A., Kakuk, L., Seddon, J., Bernstein, P., Lewis, R., Tammur, T., Yang, Z., Li, L., Zhang, H., Yashar, B., Liu, J., Petrukhin, K., Sieving, P.A., Allikmets, R. (2001). Evaluation of the ELOVL4 gene in patients with age-related macular degeneration. Ophthalmic Genetics 22, 233.
    
    Abstract: Stargardt-like macular degeneration (STGD(3)) and autosomal dominant macular degeneration (adMD) share phenotypic characters with atrophic age-related macular degeneration (AMD). Mutations in a photoreceptor cell-specific factor involved in the elongation of very long chain fatty acids (ELOVL(4)) were shown to be associated with STGD(3), adMD, and pattern dystrophy. We screened 778 patients with AMD and 551 age-matched controls to define the role of sequence variants in the ELOVL(4) gene in age-related macular degeneration. We detected three sequence variants in the non-coding region and eight variants in the coding region. No statistically significant association was observed between sequence variants in the ELOVL(4) gene and susceptibility to AMD. However, for the detection of modest effects of multiple alleles in a complex disease, the analysis of larger cohorts of patients may be required.
    
    
23. Yang, Z., Peachey, N.S., Moshfeghi, D.M., Chorich, L., Thirumalaichary, S., Shugart, Y., Fan, K., and Zhang, K. (2002). Mutations in the RPGR gene cause X-linked cone dystrophy. Human Molecular Genetics 11, 605.
    
    Abstract: X-linked cone dystrophy is a type of hereditary retinal degeneration characterized by a progressive dysfunction of the day vision or photopic (cone) system with preservation of night vision or scotopic (rod) function. The disease presents with a triad of photophobia, loss of color vision and reduced central vision. This phenotype is distinct from retinitis pigmentosa (RP) in which there are prominent night and peripheral vision disturbances. X-linked cone dystrophy is a genetically heterogeneous disorder, with linkage to loci on Xp11.4--Xp21.1 (COD1, OMIM 304020) and Xq27 (COD2, OMIM 303800). COD1 maps to a region that harbors the RPGR gene, mutations in which account for >70% of patients with X-linked RP. The majority of these mutations reside in one purine-rich exon, ORF15, encoding 567 amino acids with a repetitive domain rich in glutamic acid residues. We mapped two families with X-linked cone dystrophy to the COD1 locus and identified two distinct mutations in ORF15 in the RPGR gene (ORF15+1343_1344delGG and ORF15+694_708del15) leading to a frame-shift and premature termination of translation in one case and a deletion of five amino acids in another. Consistent with expression of RPGR in rods and cones, our results show that mutations in RPGR, in addition to X-linked RP, can also cause cone-specific degeneration. (full text pdf)
    
    
24. Yang, Z., Yu, Z., Li, Y., Jacobson, S., Thirumalaichary, S., Zack, D. J., K. Zhang, K. A novel mutation in RDS/Peripherin gene causing butterfly-shaped pattern dystrophy and adult onset foveal macular dystrophy. Submitted to Ophthalmology
    
    Abstract: OBJECTIVES: To identify the disease-causing mutation in a large family segregating dominantly inherited butterfly-shaped pattern dystrophy (BPD) and to describe the microscopic pathological changes observed in a member of this family. METHODS: Seventeen individuals at risk for dominantly inherited BPD in a family were examined and blood samples obtained. Linkage analysis and mutation screening of the human retinal degeneration slow (RDS)/peripherin locus were performed. Light and electron microscopic examinations were performed on 1 postmortem eye of 1 affected individual. RESULTS: Four individuals demonstrated macular degenerative changes with diminished visual acuity, and 3 others exhibited early signs of atrophy without visual deficits. Microscopic examination of the left eye of 1 patient revealed an area of total loss of the retinal pigment epithelium (RPE) and photoreceptor cell layer with intact choriocapillaris and lipofuscin-containing cells in the subretinal space. Outside the area of RPE atrophy, the RPE was greatly distended by lipofuscin. The disease locus in this family was mapped to 6p21.2, the region of the RDS/peripherin gene. Further analysis identified a G-->A change at nucleotide position 637 of RDS/peripherin, predicting a novel Cys213Tyr substitution in all affected members of the family. CONCLUSIONS: This study describes a new RDS/peripherin mutation for BPD and provides the first combined genetic-pathological study of this condition, to our knowledge. CLINICAL RELEVANCE: Accumulation of lipofuscin in RPE is a prominent feature of several retinal disorders, including age-related macular degeneration. Further elucidation of the cellular and molecular mechanism of BPD may provide insight into pathogenesis and lead to novel treatment approaches for this and other macular degenerations.
    
    
25. Yang, Z., Lin, W., Moshfehgi, D. M., Thirumalaichary, S., Zhang, H., Kaiser, P. K., Traboulsi, E., Zhang, K. A novel mutation in the peripherin/RDS gene causes adult-onset foveomacular dystrophy. Am J of Ophthalmology (in press)
    
    Abstract: PURPOSE: To describe a novel mutation in the RDS/Peripherin gene that results in a moderately severe form of adult-onset foveomacular dystrophy. DESIGN: Observational case series. METHODS: Selected members of a family with adult-onset foveomacular dystrophy underwent complete ophthalmic evaluation, including fundus photography and fluorescein angiography, in a tertiary care referral center. The study population consisted of 12 members of a Caucasian kindred. After providing informed consent, patients donated blood for genomic DNA extraction and mutational screening using standard techniques. The main outcome measure were the presence of a RDS/Peripherin gene mutation in a patient with the disease and its absence in unaffected family members and controls. RESULTS: Eight affected family members and no unaffected family members demonstrated a single guanine base deletion at nucleotide 112 that led to premature termination at amino acid 38 of RDS/Peripherin polypeptide. This frameshift mutation results in truncation of nearly 90% of the gene product, thus probably representing a null allele. That results in a relatively severe phenotype, with choroidal neovascularization developing in two patients and geographic atrophy involving the macula in three patients. CONCLUSIONS: We describe a frameshift null mutation in the RDS/Peripherin gene associated with a relatively severe manifestation of adult-onset foveomacular dystrophy in affected family members.
    
    
26. Kniazeva M, Sieber M, McCauley S, Zhang K , Watts JL, Han M (2003). Suppression of the ELO-2 FA Elongation Activity Results in Alterations of the Fatty Acid Composition and Multiple Physiological Defects, Including Abnormal Ultradian Rhythms, in Caenorhabditis elegans. Genetics 163, 159-69.
    
    Abstract: While the general steps of fatty acid (FA) biosynthesis are well understood, the individual enzymes involved in the elongation of long chain saturated and polyunsaturated FA (PUFA) are largely unknown. Recent research indicates that these enzymes might be of considerable physiological importance for human health. We use Caenorhabditis elegans to study FA elongation activities and associated abnormal phenotypes. In this article we report that the predicted C. elegans F11E6.5/ELO-2 is a functional enzyme with the FA elongation activity. It is responsible for the elongation of palmitic acid and is involved in PUFA biosynthesis. RNAi-mediated suppression of ELO-2 causes an accumulation of palmitate and an associated decrease in the PUFA fraction in triacylglycerides and phospholipid classes. This imbalance in the FA composition results in multiple phenotypic defects such as slow growth, small body size, reproductive defects, and changes in rhythmic behavior. ELO-2 cooperates with the previously reported ELO-1 in 20-carbon PUFA production, and at least one of the enzymes must function to provide normal growth and development in C. elegans. The presented data indicate that suppression of a single enzyme of the FA elongation machinery is enough to affect various organs and systems in worms. This effect resembles syndromic disorders in humans. (full text pdf)
    
    
27. Johnson S, Halford S, Morris AG, Patel RJ, Wilkie SE, Hardcastle AJ, Moore AT, Zhang K , Hunt DM. (2003). Genomic organisation and alternative splicing of human RIM1, a gene implicated in autosomal dominant cone-rod dystrophy (CORD7). Genomics 81, 304-14.
    
    Abstract: A mutation has been identified in the Rab3A-interacting molecule (RIM1) gene in CORD7, an autosomal dominant cone-rod dystrophy that localises to chromosome 6q14. The G to A point mutation results in an Arg844His substitution in the C(2)A domain of the protein that segregates with disease. This mutation is absent in over 200 control chromosomes, indicating that it is not a common polymorphism, and the almost complete sequence conservation of the C(2)A domain between human and rat RIM1 is consistent with a disease role for the change. RIM1 is expressed in brain and photoreceptors of the retina where it is localised to the pre-synaptic ribbons in ribbon synapses. The RIM1 gene is composed of at least 35 exons, spans 577 kb of genomic DNA, and encodes a protein of up to 1693 residues. The transcript shows extensive alternative splicing involving exons 17, 21-26 and 28-30. (full text pdf)
    
    
28. Zhang XM, Yang Z, Karan G, Hashimoto T, Baehr W, Yang XJ, Zhang K. (2003) Elovl4 mRNA distribution in the developing mouse retina and phylogenetic conservation of Elovl4 genes. Molecular Vision 9, 301.
    
    Abstract: PURPOSE: Stargardt-like macular dystrophy (STGD3) is an autosomal dominant form of early onset macular degeneration. The disease causing gene ELOVL4 encodes a protein that belongs to a family of proteins functioning in elongation of long chain fatty acids. The purpose of this study is to characterize cross-species conservation of ELOVL4 and investigate its mRNA distribution in the developing mouse eye. METHODS: Bovine and porcine orthologs of the human ELOVL4 gene were cloned using RT-PCR method. EST and HTGS databases were searched for orthologs of ELOVL4. Cross-species alignments were performed using ClustalW. In situ hybridizations using murine Elovl4 probes were performed on frozen sections of mouse eyes. RESULTS: Elovl4 orthologs from mammalian to invertebrate species share strong sequence homology with human ELOVL4 at the amino acid level, suggesting functional conservation of Elovl4 during evolution. Expression of Elovl4 in mouse retina begins at E15 during embryogenesis and persists in postnatal stages. However, Elovl4 is predominantly expressed in the retinal ganglion cells at P1-P3, followed by predominant expression in the outer nuclear layer at P7, with its final expression enriched in inner segments of photoreceptors. CONCLUSIONS: Elovl4 expression in developing retina follows a dynamic pattern. It switches from predominant ganglion cell expression in embryonic and early postnatal development to predominant expression in the photoreceptor inner segments in later stages. Phylogenetic analysis reveals strong conservation of Elovl4 among different species throughout the vertebrate subphylum consistent with our hypothesis that ELOVL4 performs a fundamentally important function. (full text pdf)
    
    
29. Karan G, Yang Z, Zhang K . (2004). Expression of wild-type and mutant ELOVL4 in cell culture: subcellular localization and cell viability, Molecular Vision 10, 248.
    
    Abstract: PURPOSE: ELOVL4 is a member of the fatty acid elongase (ELO) family of genes. Mutations of this gene are responsible for autosomal dominant Stargardt-like macular degeneration. However, the specific role of ELOVL4 in photoreceptor cells and the mechanism by which mutations in ELOVL4 causes macular degeneration are not known. In this study we examined the subcellular localization of wild type (wt) and mutant (mt) ELOVL4 EGFP fusion protein and the potential functional consequence of mtELOVL4 expression on cell viability. METHODS: Wt and mt ELOVL4 were expressed as EGFP fusion proteins in NIH 3T3 and HEK293 cells. Subcellular localizations of the fusion proteins were determined with a series of organelle-specific markers for endoplasmic reticulum (pDsRed2-ER), mitochondria (pDsRed2-Mito), peroxisomes (pDsRed2-Peroxi), and Golgi (BODIPY TR). Transfected cells were viewed using confocal and episcopic-fluorescence microscopy. Western blot analysis was performed to assess protein expression using an anti-GFP antibody. TUNEL staining was used to quantify apoptotic cell death. RESULTS: In cell transfection studies, wtELOVL4/EGFP fusion protein localized preferentially to the endoplasmic reticulum (ER) and was not found to be discernibly present in mitochondria, peroxisomes, or Golgi. In contrast, the truncated mutant fusion protein (which has no ER retention signal) appeared to be mislocalized to other compartments within transfected cells. Transfected cells expressing mtELOVL4/EGFP fusion protein exhibited induction of apoptotic cell death. CONCLUSIONS: Unlike wtELOVL4/EGFP fusion protein, the mtELOVL4/EGFP fusion protein did not localize to the ER but rather appeared to be sequestered elsewhere in an aggregated pattern in the cytoplasm. The apoptosis induced by the mutant ELOVL4 fusion protein may be the mechanism whereby photoreceptor cells degenerate in Stargardt-like macular degeneration. Our study has provided an important in vitro model system for further assessment of ELOVL4 biochemical functions. (full text pdf)
    
    
30. Toomes C, Bottomley HM, Jackson RM, Towns KV, Scott S, Mackey DA, Craig JE, Jiang L, Yang Z, Trembath R, Woodruff G, Gregory-Evans CY, Gregory-Evans K, Parker MJ, Black GCM, Downey LM, Zhang K and Inglehearn CF. (2004). Mutations in LRP5 or FZD4 underlie the common FEVR locus on chromosome 11q. Am J Hum Genetics, 74, 721.
    
    Abstract: Familial exudative vitreoretinopathy (FEVR) is an inherited blinding disorder of the retinal vascular system. Autosomal dominant FEVR is genetically heterogeneous, but its principal locus, EVR1, is on chromosome 11q13-q23. The gene encoding the Wnt receptor frizzled-4 (FZD4) was recently reported to be the EVR1 gene, but our mutation screen revealed fewer patients harboring mutations than expected. Here, we describe mutations in a second gene at the EVR1 locus, low-density-lipoprotein receptor-related protein 5 (LRP5), a Wnt coreceptor. This finding further underlines the significance of Wnt signaling in the vascularization of the eye and highlights the potential dangers of using multiple families to refine genetic intervals in gene-identification studies. (full text pdf)
    
    
31. Xu, Q, Wang, Y, Dabdoub, A, Smallwood, PM, Williams, J, Woods, C, Kelley, MW, Jiang, L, Tasman, W, Zhang, K, Nathans, J. (2004) Vascular development in the retina and inner ear: control by Norrin and Frizzled-4, a high-affinity ligand-receptor pair. Cell, 116:883.
    
    Abstract: Incomplete retinal vascularization occurs in both Norrie disease and familial exudative vitreoretinopathy (FEVR). Norrin, the protein product of the Norrie disease gene, is a secreted protein of unknown biochemical function. One form of FEVR is caused by defects in Frizzled-4 (Fz4), a presumptive Wnt receptor. We show here that Norrin and Fz4 function as a ligand-receptor pair based on (1) the similarity in vascular phenotypes caused by Norrin and Fz4 mutations in humans and mice, (2) the specificity and high affinity of Norrin-Fz4 binding, (3) the high efficiency with which Norrin induces Fz4- and Lrp-dependent activation of the classical Wnt pathway, and (4) the signaling defects displayed by disease-associated variants of Norrin and Fz4. These data define a Norrin-Fz4 signaling system that plays a central role in vascular development in the eye and ear, and they indicate that ligands unrelated to Wnts can act through Fz receptors. (full text pdf)
    
    
32. Toomes C, Bottomley HM, Scott S, Mackey DA, Craig JE, Appukuttan B, Stout JT, Zhang K , Black GCM, Fryer A, Downey LM, Inglehearn CF. Spectrum and frequency of FZD4 mutations in familial exudative vitreoretinopathy (FEVR). Invest Ophthalmol Vis Sci, in press.
    
    Abstract: PURPOSE: Mutations in the frizzled-4 gene (FZD4) have recently been associated with autosomal dominant familial exudative vitreoretinopathy (FEVR) in families linking to the EVR1 locus on the long arm of chromosome 11. The purpose of this study was to screen FZD4 in a panel of 40 patients with FEVR to identify the types and location of mutations and to calculate what proportion of this heterogeneous condition is attributable to FZD4 mutations. METHODS: PCR products were generated from genomic DNA with primers designed to amplify the coding sequence of FZD4. The PCR products were screened for mutations by single-strand conformational polymorphism-heteroduplex analysis (SSCP-HA) and by direct sequencing. RESULTS: In total, eight mutations were identified, seven of which were novel. Three were deletions (c957delG, c1498delA, and c1501-1502delCT), one was a nonsense mutation (Q505X), and four were missense mutations (G36D, M105T, M157V, and S497F). CONCLUSIONS: Eight mutations have been identified in the FZD4 gene in a cohort of 40 unrelated patients with FEVR. This result indicates that FZD4 mutations are responsible for only 20% of FEVR index cases and suggests that the other FEVR loci may account for more cases than previously anticipated. (full text pdf)
    
    
33. Yang, Z., Li, Y., Jacobson, SG, Thirumalaichary, S., Zack, D. J., Jacobson, SG , Zhang, K . A Novel RDS/Peripherin Gene Mutation Associated with Diverse Macular Phenotypes. Ophthalmic Genetics, in press.
    
    Abstract: PURPOSE: To describe a novel mutation in the RDS/Peripherin gene that results in a moderately severe form of adult-onset foveomacular dystrophy. DESIGN: Observational case series. METHODS: Selected members of a family with adult-onset foveomacular dystrophy underwent complete ophthalmic evaluation, including fundus photography and fluorescein angiography, in a tertiary care referral center. The study population consisted of 12 members of a Caucasian kindred. After providing informed consent, patients donated blood for genomic DNA extraction and mutational screening using standard techniques. The main outcome measure were the presence of a RDS/Peripherin gene mutation in a patient with the disease and its absence in unaffected family members and controls. RESULTS: Eight affected family members and no unaffected family members demonstrated a single guanine base deletion at nucleotide 112 that led to premature termination at amino acid 38 of RDS/Peripherin polypeptide. This frameshift mutation results in truncation of nearly 90% of the gene product, thus probably representing a null allele. That results in a relatively severe phenotype, with choroidal neovascularization developing in two patients and geographic atrophy involving the macula in three patients. CONCLUSIONS: We describe a frameshift null mutation in the RDS/Peripherin gene associated with a relatively severe manifestation of adult-onset foveomacular dystrophy in affected family members. (full text pdf)
    
    
34. Payne M, Yang Z, Katz BJ, Warner JEA, Weight CJ, ZhaoY, Pearson ED, Treft RL, Hillman T, Kennedy RJ, Meire FM, Zhang K. Dominant optic atrophy, sensorineural hearing loss, ptosis, and ophthalmoplegia: a syndrome caused by a missense mutation in OPA1, AJO, Accepted.
    
    
35. Berry V,Yang Z, Addison PAF, Karan G, Jiang L, Lin W, Hu J, Yang R, Moore A, Zhang K , and Bhattacharya SS. Recurrent 17bp duplication in PITX3 is primarily associated with posterior polar cataract (CPP4). J of Med. Genetics, in press.
    
    Abstract: The human lens is a unique tissue continually evolving and ever increasing in size primarily formulated by placement of newly differentiated fibre cells in concentric lamellae. This is analogous to an onion skin arrangement. This differentiation triggers loss of both nucleus and other cell organelles. Furthermore, the main factors for maintaining lens clarity are the tight and ordered mutual binding of the fibre cells and the packing of their intracellular building blocks termed crystallins. It is known that lens transparency is a crucial trade-off between the concentration of these macromolecules and their hydration. Disruption of normal levels of hydration can lead to opacity. (full text pdf)

Reviews

1. Zhang, K., Wang, M., Munier, F., Roth, D., Shields, J.A., and Donoso, L.A. (1993). Molecular genetics of retinoblastoma. International Ophthalmology Clinics 33, 53.
2. Zhang, K., Nguyen, E., Crandall, A., and Donoso, L.A. (1995). Genetic and molecular studies of macular dystrophies: recent development. Survey of Ophthalmology 40, 51.
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4. Garibaldi, D, and Zhang, K. (1999). Molecular genetics of macular degeneration. Intl Ophthal. Clinics, 39, 117.

 

 

Last updated: March 15, 2005