Developmentally regulated expression of the preproinsulin gene in the chicken embryo during gastrulation and neurulation.
Despite the absence of a pancreas, which develops between embryonic day 3 (E3) to E4, previous studies showed that insulin receptors are widely expressed in chicken embryos from the blastoderm stage (unincubated embryo, E0) through gastrulation (E0.5-E1), neurulation (E1.5-E2), and organogenesis. We now characterize prepancreatic preproinsulin gene expression and its regulation, using a highly sensitive modification of the polymerase chain reaction. We found preproinsulin messenger RNA (mRNA) expression at all stages, from the unincubated chicken blastoderm through early organogenesis, with the highest expression in embryos undergoing gastrulation. In situ hybridization analysis of E1-E1.5 embryos in toto showed widespread distribution of preproinsulin mRNA in a pattern similar to that of insulin receptor mRNA. In contrast, insulin-like growth factor-I mRNA expression appeared later than preproinsulin mRNA in the embryo; it was first demonstrable in the head portion of E3 and was found in head, trunk, and caudal regions by E4. With a novel culture system for chicken embryos during neurulation, we examined whether glucose regulated prepancreatic preproinsulin mRNA expression. Embryos cultured in glucose-free medium had increased preproinsulin mRNA with respect to the value in ovo, but the addition of 17 mM glucose had no stimulatory effect. In marked contrast, in organ cultures of E13 pancreas, insulin mRNA expression decreased in glucose-free medium by 50% relative to that in ovo. The addition of glucose restored the levels to a concentration similar to that found in ovo. Exogenous insulin added to cultured E1.5 embryos increased protein and DNA synthesis. We conclude that the preproinsulin gene is widely expressed in chicken embryo structures throughout gastrulation and neurulation. This prepancreatic preproinsulin mRNA is differentially regulated compared to the pancreatic mRNA. Preproinsulin gene products may have a role in cell proliferation, differentiation, or survival in very early avian embryos at a time when insulin-like growth factor-I expression is absent or undetectable.
Pérez-Villamil
B
B
Department of Cell and Developmental Biology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain.
de la Rosa
E J
EJ
Morales
A V
AV
de Pablo
F
F
D000818:Animals; D001483:Base Sequence; D002642:Chick Embryo / Q000378:metabolism; D002855:Chromatography, Thin Layer; D005314:Embryonic and Fetal Development; D005775:Gastrula / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental; D017403:In Situ Hybridization; D007328:Insulin; D007334:Insulin-Like Growth Factor I / Q000235:genetics; D008969:Molecular Sequence Data; D009420:Nervous System / Q000196:embryology*; D015345:Oligonucleotide Probes / Q000235:genetics; D010179:Pancreas / Q000196:embryology; D016133:Polymerase Chain Reaction; D011384:Proinsulin / Q000235:genetics*; D011498:Protein Precursors / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic
D007328:Insulin; D015345:Oligonucleotide Probes; D011498:Protein Precursors; D012333:RNA, Messenger; C013737:preproinsulin; D007334:Insulin-Like Growth Factor I; D011384:Proinsulin