The factors involved in regulating alternative splicing of the human adenylyl cyclase stimulatory G-protein G␣ s in different cell types remain undefined. We have designed a G␣ s minigene that retains the signals required for G␣ s alternative splicing in vivo. Employing transient transfection of human myometrial smooth muscle cells and HeLa cells, as well as in vitro splicing assays, we have provided evidence that the antagonistic splicing factors SF2/ASF and hnRNPA1 act as potent regulators of G␣ s isoform expression in these cells. Both SF2/ASF and hnRNPA1 control the selection of competing 5-splice sites and respectively promote inclusion or skipping of the small cassette-type exon 3 of G␣ s transcripts, resulting in the generation of G␣ s -long and G␣ sshort mRNA isoforms. We have also provided evidence that SF2/ASF and hnRNPA1 play a role in 3-splice site selection involving the use of a non-canonical TG 3-splice site preceding exon 4. Using a score-matrix analysis to identify putative exonic enhancer sequences (ESEs), we found multiple high score ESE motifs for SF2/ASF, SC35, and SRp40 in exon 3 of G␣ s . These results suggest that tissue-specific expression of SF2/ASF and hnRNPA1 governs the expression of alternative isoforms of G␣ s in these different cells types.The GTP-binding protein G␣ s is the primary stimulatory component of adenylyl cyclase in most cell types and has also been associated with activation of dihydropyridine-sensitive voltage-gated calcium channels (1) in skeletal muscle as well as inactivation of cardiac sodium channels (2). Two ubiquitously expressed forms of G␣ s have been identified via ADP-ribosylation with cholera toxin or by Western blotting (3, 4). These proteins migrate in SDS-PAGE with apparent molecular masses of 52 and 45 kDa, depending on the experimental conditions, and have been designated the long and short isoforms of G␣ s , respectively (3-5). Both isoforms of G␣ s are generated by alternative splicing of a single precursor mRNA transcript.In humans, a single copy of the G␣ s gene is found on chromosome 20 and is composed of 13 exons separated by 12 introns, altogether spanning a 20-kb region of genomic DNA (6).Cloning of the human G␣ s gene and G␣ s complementary DNAs showed that the short form of G␣ s is distinguished from the long form by the exclusion of the 45-bp exon 3, which encodes 15 amino acids. Similar studies have pointed to the existence of two additional G␣ s mRNA species. These isoforms may be produced by the use of an unusual alternative TG 3Ј-splice site, instead of the consensus AG 3Ј-splice site upstream of exon 4, resulting in the inclusion of an extra triplet coding for a serine residue after amino acids 87 and 72 of the long and short forms of G␣ s , respectively (6) (Fig. 1). It has been suggested that inclusion of this extra serine residue into G␣ s proteins confers additional consensus sequence sites for phosphoregulation by protein kinases C and A (7, 8). Tissue-dependent alternative splicing of the G␣ s precursor transcript may therefore r...

Summary A case of phaeochromocytoma with marked transient elevation of creatine kinase levels is presented. No obvious cause for the elevation was found in life, but on autopsy a non-specific focal myositis was discovered. Possible reasons for the raised creatine kinase levels and focal myositis are discussed.

There is now extensive evidence to indicate that components of the cAMP signaling pathway are up-regulated in the human myometrium during pregnancy so as to potentiate the maintenance of uterine quiescence until term. In many tissue and cell types, increased signaling of the cAMP pathway results in profound changes in gene expression that are catalyzed via stimulation of PKA and activation of cAMP-dependent transcription factors that bind cAMP response elements (CREs) within the promoter regions of affected genes. In the myometrium, these CRE containing genes include beta2-adrenoceptor, cyclo-oxygenase 2, oxytocin receptor, and connexin-43. In preliminary investigations, we reported the differential expression of members of the cAMP bZIP protein family in the myometrium during pregnancy and labor. In this present study, we have now identified and functionally characterized these proteins with respect to myometrial gene expression. We report the identification of a 39,000 mol wt CRE response element modulator protein (CREM)tau2alpha protein having both transactivation and transrepressor properties whose expression is sequentially decreased in the myometrium during gestation and parturition. In contrast, expression of a myometrial 28,000 mol wt CREMalpha protein having only transrepressor actions progressively increased in the myometrium during pregnancy and labor. Similarly, we have isolated two ATF2 proteins of 60,000 and 28,000 mol wts, which represent full-length ATF2 and a novel small isoform of ATF2 that we have termed ATF2-small (ATF2-sm). These proteins are potent transactivators of gene expression and appear to be spatially expressed within the myometrium of the upper and lower uterine regions. The identification and functional characterization of these basic region/leucine zipper proteins in the myometrium may provide further insight into the molecular mechanisms regulating uterine activity during fetal maturation and parturition.