ABSTRACT
Agonist-induced desensitization of adenosine A1 receptor-mediated inhibition of adenylyl cyclase has been studied in cerebellar granule cells. Exposure of cells to the adenosine A1 receptor agonist R-phenylisopropyl adenosine (R-PIA) from 2 to 48 h brings about desensitization of this signal transduction pathway. Associated with the desensitization process, a decrease in radioligand binding performed in intact cells with the adenosine A1 receptor agonist [3H]cyclohexyladenosine (CHA) has been detected. Simultaneously, an increase of adenosine A1 radioligand binding has also been detected in microsomes. A decrease in the steady-state level of alpha-Gi in both, plasma membrane and microsomes also has been detected during the desensitization process. These data may account for the desensitization of the inhibitory pathway of the adenylyl cyclase in cerebellar granule cells described herein. After a transient increase in adenosine A1 receptor mRNA, no changes were observed in this parameter after 12 hr of treatment with the adenosine A1 agonist R-PIA, suggesting a post-transcriptional regulation of this receptor during long-term desensitization.
Subject(s)
Adenylyl Cyclase Inhibitors , Cerebellum/cytology , Cerebellum/enzymology , Purinergic P1 Receptor Agonists , Adenosine/analogs & derivatives , Adenosine/pharmacology , Animals , Cells, Cultured , Cerebellum/drug effects , GTP-Binding Proteins/metabolism , RNA, Messenger/drug effects , Rats , Rats, Sprague-Dawley , Receptors, Purinergic P1/genetics , Receptors, Purinergic P1/metabolism , Signal Transduction/drug effectsABSTRACT
Replacement-variant H3.3 histones have been isolated and sequenced in different eukaryotes, but no functional H3.3A gene has been characterized in the mouse so far. We have cloned an H3.3A cDNA from a mouse fetal ovary library, differentially screened with testis versus somatic cDNA probes. We showed this gene contains a region homologous to the reverse and complementary alpha-globin gene. We believe such a structure could have been generated by retroposition during the evolution of both globin and histone gene families. The sequence coding for H3.3A is 76.6% homologous to the mouse H3.3B gene at the nucleotide level and differs in only one amino acid at the protein level. The high degree of homology between these genes and the H3.3 variant histones from other eukaryotes reveals the conservation of these replication-independent class of histones throughout evolution. Analysis of gene expression reveals a developmental regulation concurrent with meiotic progression, with the highest level of transcript detection coincident with meiotic onset during both oogenesis and spermatogenesis.
Subject(s)
Gametogenesis/genetics , Gene Expression Regulation, Developmental , Globins/genetics , Histones/genetics , Meiosis/genetics , Amino Acid Sequence , Animals , Base Sequence , Female , Male , Mice , Molecular Sequence Data , RNA, Messenger/analysis , RNA, Messenger/genetics , Sequence Analysis, DNA , Sequence Homology, Nucleic AcidABSTRACT
Cloning and characterization of H3.3A variant histone expression has recently been reported to be associated with meiotic development in mouse testis and ovary. Using Northern analysis and in situ hybridization, the pattern of H3.3A expression was studied during the development of different tissues. In addition to the differential expression detected in male and female meiosis, H3.3A was found to be highly expressed in preantral follicles of adult ovaries and in the basal regions of seminiferous epithelium corresponding to spermatogonia. Different patterns of expression were observed in somatic tissues, which also differed with respect to the developmental stage of the tissue. The lowest expression was detected in adult skeletal muscle. High expressions were found in foetal liver and spinal cord. These different expressions might reflect a possible function of H3.3A in cell differentiation as detected in MEL cells.
Subject(s)
Gene Expression Regulation, Developmental , Histones/genetics , Ovary/embryology , RNA, Messenger/analysis , Testis/embryology , Animals , Blotting, Northern , Cell Differentiation , Cloning, Molecular , Female , Histocytochemistry , Image Processing, Computer-Assisted , In Situ Hybridization , Liver/embryology , Liver/metabolism , Male , Mice , Molecular Sequence Data , Organ Specificity , Ovary/metabolism , RNA, Messenger/genetics , Seminiferous Tubules/cytology , Seminiferous Tubules/embryology , Seminiferous Tubules/metabolism , Spinal Cord/embryology , Spinal Cord/metabolism , Testis/metabolismABSTRACT
Accumulation of transcripts from N-ras and unr genes was comparatively analyzed during the development of germline and somatic tissues. Northern blots on fetal and postnatal samples from somatic tissues, including brain, skeletal muscle, liver, kidney, small intestine and heart were studied together with ovaries and testis. While the expression of N-ras was rather stable all along the development of the different tissues analyzed, the expression of unr exhibited a specific pattern in some tissues. Specifically, in testis, there is a developmental regulation of the relative accumulation of the three alternative transcripts. Unr has a relative high expression in testes and heart but the accumulation seems to be different for the different size transcripts in each case. However, the expression in small intestine is practically absent in adults. From the comparative analysis of the expression of both genes, N-ras and unr, we propose that the regulation of N-ras is not directly coordinated with unr expression during the development. However, the expression of unr and its alternative transcripts is developmentally and differentially regulated in small intestine, heart and testis. The change in the pattern of accumulation during testis development from long to small alternative transcripts, could be interpreted in terms of possible alleviation of transcription interference of N-ras.
Subject(s)
Genes, ras , Ovary/embryology , Testis/embryology , Aging , Animals , Blotting, Northern , Female , Gene Expression , Heart/embryology , Intestine, Small/embryology , Intestine, Small/ultrastructure , Male , Mice , Mice, Inbred Strains , Myocardium/ultrastructure , Ovary/ultrastructure , Testis/ultrastructureABSTRACT
Isolation and initial characterization of clones from a mouse fetal ovary cDNA library are reported. In mammals, the development of female gametogenesis, including the prophase of the first meiotic division, begins in the fetus. A parallel conserved process occurs in males, from puberty onwards. We cloned cDNAs preferentially expressed in both sexes during gametogenesis considering both the meiotic programme and the non-germ cells participating in the developmental regulation of gametogenesis. From 183 clones that showed positive hybridization with adult testis and negligible hybridization with somatic probes, 32 clones were selected. After partial sequencing, 16 were found to correspond to known genes that are expressed during other programmes of differentiation and development. Sixteen clones represent novel sequences. The accumulation of corresponding transcripts was analysed by developmental northern blots during spermatogenesis, and genes expressed during gametogenesis were characterized together with others showing pleiotropic expression in different tissues. Additional specific, gametic transcripts were also detected.
