Green fluorescent protein expression in germ-line transmitted transgenic zebrafish under a stratified epithelial promoter from keratin8.
A zebrafish cDNA encoding a novel keratin protein was characterized and named keratin8, or krt8. krt8 expression was initiated at 4.5 hr postfertilization, immediately after the time of zygotic genome activation. The expression is limited to a single layer of envelope cells on the surface of embryos and, in later stages, it also appears in the innermost epithelial layer of the anterior- and posteriormost portions of the digestive tract. In adult, its expression was limited to the surface layer of stratified epithelial tissues, including skin epidermis and epithelia of mouth, pharynx, esophagus, and rectum but not in the gastral and intestinal epithelia. By using a 2.2-kb promoter from krt8, several stable green fluorescent protein (gfp) transgenic zebrafish lines were established. All of these transgenic lines displayed GFP expression in tissues mentioned above except for the rectum; therefore, the pattern of transgenic GFP expression is essentially identical to that of the endogenous krt8 mRNAs. krt8-GFP fusion protein was also expressed in zebrafish embryos under a ubiquitous promoter, and the fusion protein was capable of assembling into intermediate filaments only in the epithelia that normally expressed krt8 mRNAs, indicating the specificity of keratin assembly in vivo.
Gong
Zhiyuan
Z
Department of Biological Sciences, National University of Singapore, Singapore. dbsgzy@nus.edu.sg
Ju
Bensheng
B
Wang
Xukun
X
He
Jiangyan
J
Wan
Haiyan
H
Sudha
Putter Mudumana
PM
Yan
Tie
T
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics; D004064:Digestive System / Q000196:embryology / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000378:metabolism; D004817:Epidermis / Q000196:embryology / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism*; D005260:Female; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D005813:Genes, Synthetic; D049452:Green Fluorescent Proteins; D053533:Keratin-8; D007633:Keratins / Q000096:biosynthesis* / Q000235:genetics; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D008969:Molecular Sequence Data; D009928:Organ Specificity; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014162:Transfection; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins / Q000096:biosynthesis* / Q000235:genetics
D018076:DNA, Complementary; D004274:DNA, Recombinant; D053533:Keratin-8; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D029961:Zebrafish Proteins; C486596:krt8 protein, zebrafish; D049452:Green Fluorescent Proteins; D007633:Keratins