Conservation of mouse alpha A-crystallin promoter activity in chicken lens epithelial cells.
Previous transfection experiments have shown that 162 base pairs (bp) of the 5' flanking sequence of the chicken alpha A-crystallin gene are required for promoter activity in primary chicken lens epithelial cells (PLE), while only 111 bp of the 5' flanking sequence are needed for activity of the mouse alpha A-crystallin promoter in transfected chicken PLE cells or in a SV40 T-antigen-transformed transfected mouse lens epithelial cell line (alpha TN4-1). The effect of site-directed mutations covering positions -111 to -34 of the mouse alpha A-crystallin promoter fused to the bacterial chloramphenicol acetyltransferase (CAT) gene was compared in transfected chicken PLE cells and mouse alpha TN4-1 cells; selected mutations were also examined in a nontransformed rabbit lens epithelial cell line (N/N1003A). In general, the same mutations reduced promoter activity in the transfected lens cells from all three species, although differences were noted. The mutations severely affected regions -111/-106 and -69/-40 regions in all the transfected cells examined; by contrast, mutations at positions -105/-99 and -87/-70 had a somewhat greater effect in the chicken PLE than the mouse alpha TN4-1 cells, while mutations of the -93/-88 sequence reduced expression in the alpha TN4-1 but not the PLE cells. A partial cDNA with sequence similarity to alpha A-CRYPB1 of the mouse has been isolated from a chicken lens library; mouse alpha A-CRYBP1 is a putative transcription factor which binds to the -66/-55 sequence of the mouse alpha A-crystallin promoter.(ABSTRACT TRUNCATED AT 250 WORDS)
Donovan
D M
DM
Laboratory of Molecular and Developmental Biology, NEI, NIH, Bethesda, MD 20892.
Sax
C M
CM
Klement
J F
JF
Li
X
X
Chepelinsky
A B
AB
Piatigorsky
J
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002478:Cells, Cultured; D002642:Chick Embryo; D003001:Cloning, Molecular; D003459:Crystallins / Q000737:chemistry / Q000235:genetics*; D005786:Gene Expression Regulation / Q000235:genetics*; D007908:Lens, Crystalline / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics*; D014162:Transfection
D003459:Crystallins; D011993:Recombinant Fusion Proteins