Identification and developmental expression of a smooth-muscle gamma-actin in postmeiotic male germ cells of mice.

Mouse testis contains two size classes of actin mRNAs of 2.1 and 1.5 kilobases (kb). The 2.1-kb actin mRNA codes for cytoplasmic beta- and gamma-actin and is found throughout spermatogenesis, while the 1.5-kb actin mRNA is first detected in postmeiotic cells. Here we identify the testicular postmeiotic actin encoded by the 1.5-kb mRNA as a smooth-muscle gamma-actin (SMGA) and present its cDNA sequence. The amino acid sequence deduced from the postmeiotic actin cDNA sequence was nearly identical to that of a chicken gizzard SMGA, with one amino acid replacement at amino acid 359, where glutamine was substituted for proline. The nucleotide sequence of the untranslated region of the SMGA differed substantially from those of other isotypes of mammalian actins. By using the 3' untranslated region of the testicular SMGA, a highly specific probe was obtained. The 1.5-kb mRNA was detected in RNA from mouse aorta, small intestine, and uterus, but not in RNA isolated from mouse brain, heart, and spleen. Testicular SMGA mRNA was first detected and increased substantially in amount during spermiogenesis in the germ cells, in contrast to the decrease of the cytoplasmic beta- and gamma-actin mRNAs towards the end of spermatogenesis. Testicular SMGA mRNA was present in the polysome fractions, indicating that it was translated. These studies demonstrate the existence of an SMGA in male haploid germ cells. The implications of the existence of an SMGA in male germ cells are discussed.     

Tissue- and developmental stage-specific expression of the rat ornithine carbamoyltransferase gene in transgenic mice

Rat ornithine carbamoyltransferase (OCT; EC 2.1.3.3) is encoded by a large gene of 75 kilobases. Expression of this gene is restricted to the liver and small intestine, and there is an increase in expression late in gestation. The recombinant gene carrying 1.3 kilobases of the 5' flanking region of the gene fused to the rat OCT cDNA was microinjected into fertilized eggs, and 17 transgenic mice were produced. Expression in the liver of the transgene was detected in three mice. In these mice, the transgene expression was observed exclusively in the liver and small intestine. Expression of the transgene in the intestine was comparable to that of the endogenous mouse OCT gene, whereas expression in the liver was much lower than that of the endogenous gene. The developmental pattern of expression of the transgene was similar to that of the endogenous gene. Therefore, the 5' flanking sequence of the rat OCT gene seems to be sufficient for the developmental and tissue-specific expression of the gene. An explanation for low expression in the liver remains the subject of ongoing study.     

GABA transporter 1 transcriptional starting site exhibiting tissue specific difference

INTRODUCTIONIn the vertebrate central nervous system (CNS),GABA transporters (GAT) are believed to play animportant role in termination of GABAergiC transInission. GATI was the first cloned member of neurotransmitter transporters superfanilly[1], and soon,other three subtypes (GAT2-4) were subsequentlycloned. Since GABA is the predominant inhibitoryneurotranslliltter in CNS, abnormallty of GATs hasa direct relationship with certain kinds of nervousdisorders, such as epilepsy a…     

Cloning and functional characterization of the human GLUT7 isoform SLC2A7 from the small intestine

Facilitated glucose transporters (GLUTs) mediate transport of sugars across cell membranes by using the chemical gradient of sugars as the driving force. Improved cloning techniques and database analyses have expanded this family of proteins to a total of 14 putative members. In this work a novel hexose transporter isoform, GLUT7, has been cloned from a human intestinal cDNA library by using a PCR-based strategy (GenBank accession no. AY571960). The encoded protein is comprised of 524 amino acid residues and shares 68% similarity and 53% identity with GLUT5, its most closely related isoform. When GLUT7 was expressed in Xenopus oocytes, it showed high-affinity transport for glucose (K(m) = 0.3 mM) and fructose (IC(50) = 0.060 mM). Galactose, 2-deoxy-d-glucose, and xylose were not transported. Uptake of 100 microM d-glucose was not inhibited by 200 microM phloretin or 100 microM cytochalasin B. Northern blotting indicated that the mRNA for GLUT7 is present in the human small intestine, colon, testis, and prostate. Western blotting and immunohistochemistry of rat tissues with an antibody raised against the predicted COOH-terminal sequence confirmed expression of the protein in the small intestine and indicated that the transporter is predominantly expressed in the enterocytes' brush-border membrane. The unusual substrate specificity and close sequence identity with GLUT5 suggest that GLUT7 represents an intermediate between class II GLUTs and the class I member GLUT2. Comparison between these proteins may provide key information as to the structural determinants for the recognition of fructose as a substrate.