Isolation and characterization of the human CP49 gene promoter.

PURPOSE This study focuses on the identification of regulatory elements that contribute to lens-specific expression of the human CP49 gene within the 5'-flanking DNA sequences. METHODS The DNA sequence upstream of the human CP49 coding region was subcloned as a set of 5' and 3' deletion series. The constructs were transfected into lens (N/N1003A) and nonlens (NIH3T3) cell lines and chicken primary lens cultures, to test for promoter activity and specificity. To further test the specificity, a portion of the 5' flanking DNA sequence was used to drive transgene expression in mice. The flanking DNA sequence was analyzed for potential transcription factor-binding sites. RESULTS The 5'-flanking DNA preferentially activated reporter gene expression in a lens-preferred manner when transfected into cultured cells. Transgene expression driven by the CP49 promoter region was lens specific. Analysis of the proximal promoter sequence revealed the presence of potential binding sites for the AP-1, AP-2, and OCT-1 transcription factors and the absence of TATA and CAAT boxes. CONCLUSIONS The sequence upstream of the CP49 gene possesses promoter activity and is able to drive lens-preferred expression in both transfection and transgenic experiments. Promoter activity is dependent on the presence of the proximal 300 bp directly upstream of the coding region.

[1]  H. Nijveen,et al.  Regulatory Elements in the Rat βB2-crystallin Promoter , 2001 .

[2]  J. Hess,et al.  Development- and differentiation-dependent reorganization of intermediate filaments in fiber cells. , 2001, Investigative ophthalmology & visual science.

[3]  H. Nijveen,et al.  Regulatory elements in the rat betaB2-crystallin promoter. , 2001, Experimental eye research.

[4]  J. Hess,et al.  Autosomal-dominant congenital cataract associated with a deletion mutation in the human beaded filament protein gene BFSP2. , 2000, American journal of human genetics.

[5]  D. Weeks,et al.  A juvenile-onset, progressive cataract locus on chromosome 3q21-q22 is associated with a missense mutation in the beaded filament structural protein-2. , 2000, American journal of human genetics.

[6]  G. Barsh,et al.  Regulation of mouse lens fiber cell development and differentiation by the Maf gene. , 2000, Development.

[7]  K. Yasuda,et al.  Regulation of Lens Fiber Cell Differentiation by Transcription Factor c-Maf* , 1999, The Journal of Biological Chemistry.

[8]  Tiansen Li,et al.  Requirement for the c-Maf transcription factor in crystallin gene regulation and lens development. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[9]  A. Cvekl,et al.  Dual Roles for Pax-6: a Transcriptional Repressor of Lens Fiber Cell-Specific β-Crystallin Genes , 1998, Molecular and Cellular Biology.

[10]  H. Kondoh,et al.  Identification and functional analysis of the mouse lens filensin gene promoter. , 1998, Gene.

[11]  J. Hess,et al.  Primary sequence, secondary structure, gene structure, and assembly properties suggests that the lens-specific cytoskeletal protein filensin represents a novel class of intermediate filament protein. , 1998, Experimental eye research.

[12]  H. Ogino,et al.  Induction of lens differentiation by activation of a bZIP transcription factor, L-Maf. , 1998, Science.

[13]  R. Lovell-Badge,et al.  Sox1 directly regulates the gamma-crystallin genes and is essential for lens development in mice. , 1998, Genes & development.

[14]  R. Quinlan,et al.  Lens intermediate filament proteins. , 1998, Sub-cellular biochemistry.

[15]  H. Ge,et al.  Overlapping Sp1 and AP2 binding sites in a promoter element of the lens-specific MIP gene. , 1998, Nucleic acids research.

[16]  P. Lichter,et al.  The mouse filensin gene: structure and evolutionary relation to other intermediate filament genes , 1997, FEBS letters.

[17]  T. Sun,et al.  Regulation of K3 keratin gene transcription by Sp1 and AP-2 in differentiating rabbit corneal epithelial cells , 1997, Molecular and cellular biology.

[18]  A. Cvekl,et al.  Lens development and crystallin gene expression: many roles for Pax‐6 , 1996, BioEssays : news and reviews in molecular, cellular and developmental biology.

[19]  M. Pisano,et al.  Isolation and characterization of the 5'-flanking sequence of the human ocular lens MIP gene. , 1995, Gene.

[20]  R. Lovell-Badge,et al.  Involvement of SOX proteins in lens‐specific activation of crystallin genes. , 1995, The EMBO journal.

[21]  M. Kaufman,et al.  Craniofacial abnormalities in homozygous Small eye (Sey/Sey) embryos and newborn mice. , 1995, Journal of anatomy.

[22]  R. Pankov,et al.  AP-1, ETS, and transcriptional silencers regulate retinoic acid-dependent induction of keratin 18 in embryonic cells , 1994, Molecular and cellular biology.

[23]  L. Hu,et al.  Activation of keratin 19 gene expression by a 3' enhancer containing an AP1 site. , 1994, The Journal of biological chemistry.

[24]  H. Kondoh,et al.  Overlapping positive and negative regulatory elements determine lens-specific activity of the delta 1-crystallin enhancer , 1993, Molecular and cellular biology.

[25]  J. Hess,et al.  cDNA analysis of the 49 kDa lens fiber cell cytoskeletal protein: a new, lens-specific member of the intermediate filament family? , 1993, Current eye research.

[26]  T. Watanabe,et al.  cDNA sequence analysis of CP94: rat lens fiber cell beaded-filament structural protein shows homology to cytokeratins. , 1992, Biochemical and biophysical research communications.

[27]  E. Fuchs,et al.  Transcription factor AP2 and its role in epidermal-specific gene expression. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[28]  S. Chung,et al.  Human αB-crystallin gene and preferential promoter function in lens , 1990 .

[29]  J. Piatigorsky,et al.  Human alpha B-crystallin gene and preferential promoter function in lens. , 1990, Genomics.

[30]  E. Fuchs,et al.  Isolation, sequence, and expression of a human keratin K5 gene: transcriptional regulation of keratins and insights into pairwise control , 1989, Molecular and cellular biology.

[31]  R. Fisher.  The Ocular Lens: Structure, Function, and Pathology , 1986 .

[32]  J. Piatigorsky,et al.  Lens-specific expression and developmental regulation of the bacterial chloramphenicol acetyltransferase gene driven by the murine alpha A-crystallin promoter in transgenic mice. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[33]  L. Tsui,et al.  Lens-specific promoter activity of a mouse gamma-crystallin gene. , 1985, Molecular and cellular biology.

[34]  J. Piatigorsky,et al.  Lens-specific expression of the chloramphenicol acetyltransferase gene promoted by 5' flanking sequences of the murine alpha A-crystallin gene in explanted chicken lens epithelia. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[35]  A. S. Menko,et al.  Chicken embryo lens cultures mimic differentiation in the lens. , 1984, Developmental biology.