Identification and Functional Analysis of Splice Variants of the Germ Cell Soluble Adenylyl Cyclase

Abstract

In mammalian germ cells, cAMP signaling is dependent on two forms of adenylyl cyclase, the conventional membrane-bound ACIII and a soluble form of adenylyl cyclase (sAC). Recent elucidation of the sAC sequence indicates that this enzyme is phylogenetically distinct from the membrane-bound AC, does not interact with G proteins, and its activity is regulated by bicarbonate ions. Here we have investigated the properties and regulation of this enzyme during spermatogenesis. Two different transcripts encoding a full-length and truncated sAC were identified by reverse transcriptase-polymerase chain reaction and RNase protection analysis. The truncated sAC transcript lacks exon 11 with a premature termination of the open reading frame after the catalytic domain. Reverse transcriptase-polymerase chain reaction with testis RNA from adult mouse and rat of different ages, as well as RNase protection, showed that both transcripts are absent at 11 days of age, appear between 20 and 30 days of age, and are retained in the adult testis. The presence of corresponding proteins in testis, germ cells, and spermatozoa was demonstrated by fast protein liquid chromatography and differential immunoprecipitation with full-length sAC-specific antibodies. Bicarbonate ions activated both sAC forms and increased cAMP levels in germ cells isolated from 25- and 50-day-old rats and adult rats in a concentration-dependent manner. These findings provide evidence that full-length and truncated sAC are generated by alternate splicing. Both forms are active in spermatids, and the bicarbonate present in the seminiferous tubule may be a signal that regulates cAMP levels in these cells. In mammalian germ cells, cAMP signaling is dependent on two forms of adenylyl cyclase, the conventional membrane-bound ACIII and a soluble form of adenylyl cyclase (sAC). Recent elucidation of the sAC sequence indicates that this enzyme is phylogenetically distinct from the membrane-bound AC, does not interact with G proteins, and its activity is regulated by bicarbonate ions. Here we have investigated the properties and regulation of this enzyme during spermatogenesis. Two different transcripts encoding a full-length and truncated sAC were identified by reverse transcriptase-polymerase chain reaction and RNase protection analysis. The truncated sAC transcript lacks exon 11 with a premature termination of the open reading frame after the catalytic domain. Reverse transcriptase-polymerase chain reaction with testis RNA from adult mouse and rat of different ages, as well as RNase protection, showed that both transcripts are absent at 11 days of age, appear between 20 and 30 days of age, and are retained in the adult testis. The presence of corresponding proteins in testis, germ cells, and spermatozoa was demonstrated by fast protein liquid chromatography and differential immunoprecipitation with full-length sAC-specific antibodies. Bicarbonate ions activated both sAC forms and increased cAMP levels in germ cells isolated from 25- and 50-day-old rats and adult rats in a concentration-dependent manner. These findings provide evidence that full-length and truncated sAC are generated by alternate splicing. Both forms are active in spermatids, and the bicarbonate present in the seminiferous tubule may be a signal that regulates cAMP levels in these cells. cAMP-response element-binding protein cyclic AMP-response element modulator soluble form of adenylyl cyclase adenylyl cyclase full-length sAC truncated sAC rat sAC reverse transcriptase-polymerase chain reaction fast protein liquid chromatography base pair kilobase pair Dulbecco's modified Eagle's medium radioimmunoassay ribonuclease protection assay Cyclic AMP, the product of adenylyl cyclase, is an important second messenger controlling signaling in a wide variety of cells in almost all prokaryotic and eukaryotic organisms (1Sunahara R.K. Dessauer C.W. Gilman A.G. Annu. Rev. Pharmacol. Toxicol. 1996; 36: 461-480Crossref PubMed Scopus (732) Google Scholar, 2Taussig R. Gilman A.G. J. Biol. Chem. 1995; 270: 1-4Abstract Full Text Full Text PDF PubMed Scopus (431) Google Scholar). This also holds true for the male gonad, where cAMP signaling is essential for the hormonal regulation of somatic cell differentiation and function. Similarly, acquisition of motility, capacitation, and acrosome reaction, processes that render mature spermatozoa competent to fertilize an egg, are dependent on cAMP signaling (3Garbers D.L. Tubb D.J. Kopf G.S. Biol. Reprod. 1980; 22: 526-532Crossref PubMed Scopus (21) Google Scholar, 4Visconti P.E. Stewart-Savage J. Blasco A. Battaglia L. Miranda P. Kopf G.S. Tezon J.G. Biol. Reprod. 1999; 61: 76-84Crossref PubMed Scopus (183) Google Scholar, 5Yanagimachi R. Knobil E. Neill J. The Physiology of Reproduction. Raven Press, Ltd., New York1994: 189-317Google Scholar, 6Jaiswal B.S. Eisenbach M. Hardy D.M. Garbers D.L. Fertilization. Academic Press, San Diego2001Google Scholar). Conversely, little is known about the role of the cAMP pathway during the differentiation of germ cells. That cAMP signaling may be important for the spermatogenic cell is suggested by the genetic ablation of the CREB1-related protein CREM that is expressed at a high level in differentiating spermatids (7Delmas V. van der Hoorn F. Mellstrom B. Jegou B. Sassone-Corsi P. Mol. Endocrinol. 1993; 7: 1502-1514Crossref PubMed Scopus (187) Google Scholar). Mice deficient in CREM are sterile, and spermatogenesis is arrested at the beginning of spermatid differentiation (8Nantel F. Monaco L. Foulkes N.S. Masquilier D. LeMeur M. Henriksen K. A. M. Sassone-Corsi P. 1996; PubMed Scopus Google Scholar, E. 1996; PubMed Scopus Google Scholar). this and the that activated during differentiation cAMP-response in Sassone-Corsi P. Biol. Reprod. PubMed Scopus Google Scholar, Sassone-Corsi P. A. 1996; PubMed Scopus Google that CREM as a to the genetic spermatid differentiation P. Biol. PubMed Scopus Google Scholar). CREM is and activated by the protein in a to the cAMP-response element-binding protein (7Delmas V. van der Hoorn F. Mellstrom B. Jegou B. Sassone-Corsi P. Mol. Endocrinol. 1993; 7: 1502-1514Crossref PubMed Scopus (187) Google Scholar, J. J. Sassone-Corsi P. J. 1993; PubMed Scopus Google that cAMP signaling is for male germ cells and mature cAMP is by at two a G adenylyl cyclase, is also expressed at high levels in the and in M. M. PubMed Google Scholar, M. Jegou B. J. PubMed Scopus Google Scholar). In known that a G and adenylyl cyclase is present in germ cells and spermatozoa A. PubMed Scopus Google Scholar, Cyclic Google Scholar, M. R. V. 1980; PubMed Scopus Google Scholar, K. V. J. PubMed Scopus (21) Google of the soluble form of adenylyl cyclase was by protein and J. L. A. 1999; PubMed Scopus Google Scholar). This adenylyl cyclase is an enzyme with and is to in J. L. A. 1999; PubMed Scopus Google Scholar). and not sAC activity J. L. A. 1999; PubMed Scopus Google a with the properties for the germ cell soluble cyclase M. R. V. 1980; PubMed Scopus Google Scholar, V. Mol. Endocrinol. PubMed Scopus Google Scholar, D. V. J. Endocrinol. PubMed Scopus Google Scholar). spermatozoa from all an adenylyl cyclase that is to bicarbonate A. J. Biol. Chem. Full Text PDF PubMed Google Scholar, P. J. 1993; PubMed Scopus Google Scholar). that sAC to the cyclase to bicarbonate in testis, in and in as a bicarbonate V. J. PubMed Scopus Google of the cyclase identified a protein with a of V. Mol. Endocrinol. PubMed Scopus Google Scholar, D. V. J. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google Scholar). the of the sAC an open reading frame that for a protein J. L. A. 1999; PubMed Scopus Google Scholar). these that the protein is an that is to the active form J. L. A. 1999; PubMed Scopus Google Scholar). In of this of a truncated sAC the catalytic a active protein of J. L. A. 1999; PubMed Scopus Google the present we have this and provide evidence for an for the of the sAC we that two alternate transcripts are expressed in germ cells, and both the full-length and truncated to the germ cell adenylyl cyclase the high of bicarbonate in the seminiferous tubule in germ cells we that sAC is active on during germ cell differentiation and its bicarbonate regulation may provide a by germ cell is regulated by the evidence that two distinct transcripts encoding full-length and truncated as well as the corresponding proteins, are present in mouse and rat germ cells. expressed in a these proteins have distinct In we that the full-length sAC is active and to the of cAMP levels in germ cells and in mature the high of bicarbonate ions in the seminiferous tubule we that this cAMP not in mature spermatozoa also in differentiating of the properties of the soluble testis adenylyl cyclase that this activity in a of in rat testis V. Mol. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google in D. L. J. J. Biol. Chem. Full Text PDF PubMed Google and in testis D. V. J. Endocrinol. PubMed Scopus Google Scholar, L. V. PubMed Scopus Google Scholar). the of the sAC and the that the protein by this is that the form J. L. A. 1999; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). D. L. J. J. Biol. Chem. Full Text PDF PubMed Google have that of spermatozoa a soluble cyclase with a of properties of this product were not to of the to a soluble cyclase that is activated by bicarbonate have generated K. PubMed Scopus Google provide evidence that an may the a for sAC with an exon isolated during the of a rat testis J. L. A. 1999; PubMed Scopus Google by an with rat and mouse testis we have demonstrated that this is in a of of The high with these were that is not a a and RNase protection of RNA from of mouse and rat at different of that transcripts the for of the sAC that these transcripts are at be that of the sAC protein is is the truncated the that is active that of the adenylyl cyclase activity in the testis is to is to This on the of the two forms is with the on both the sAC activity from germ cells and the of testis. an that the full-length we that is not expressed in the testis of germ cells, and its is spermatogenesis to the spermatid provide evidence for an for the of full-length and truncated the that also the form as suggested by J. L. A. 1999; PubMed Scopus Google we that this for the presence of the two sAC of evidence that the is active in germ cells. cyclase activity was after of the full-length after immunoprecipitation with an that Both and are by the of chromatography of evidence for the presence of an active sAC form with a be that this of the presence of two distinct active is with V. Mol. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google Scholar, D. L. J. J. Biol. Chem. Full Text PDF PubMed Google Scholar). cAMP levels in cells were increased the was in as a with an of with little activity in the This with the distinct properties of the two the that the in the is generated by of The that as a the as an of two as a with an This to the in properties between the and the be that we the that and as and that immunoprecipitation with an that the also the active that the full-length sAC is and activated during the assay the presence of a is with the this provide evidence that both sAC are active in germ cells, the to be that as an for In to with the catalytic we that may be regulated by is that the of for and in the of These the full-length not the truncated protein by a in of the The in activity of the the is with this spermatozoa were as a of about of the soluble activity be that the full-length sAC is retained during the differentiation of spermatids and in a with the by V. J. PubMed Scopus Google Scholar). This the that both full-length and truncated sAC a role in the and of the generated that sAC is on during and that two active sAC proteins are present at in spermatids and This that sAC are not to the of cAMP levels in mature spermatozoa that sAC is active in cAMP during germ cell This is by the that bicarbonate cAMP levels in germ cells from 25- and 50-day-old rat testis, at a of mature spermatozoa are that seminiferous tubule a distinct from that of the present at levels in In the seminiferous tubule bicarbonate between 20 and J. Reprod. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. Endocrinol. PubMed Scopus Google Scholar, J. J. PubMed Scopus Google is that in the These are to sAC in a in germ cells as for cAMP In as activity in the testis, and in spermatozoa M. E. PubMed Scopus Google Scholar, K. PubMed Scopus Google Scholar, P. J. Mol. Reprod. 1999; PubMed Scopus Google Scholar, K. J. Biol. Reprod. 1999; 61: PubMed Scopus Google Scholar, E. L. PubMed Google Scholar). and rat germ cells transcripts P. J. Mol. Reprod. 1999; PubMed Scopus Google and high levels of activity have in cells of M. E. PubMed Scopus Google E. L. PubMed Google Scholar). all these and that bicarbonate cAMP in isolated germ cells, that bicarbonate cAMP during spermatid is not known bicarbonate during the seminiferous tubule we that bicarbonate in the seminiferous tubule is of an to cAMP for spermatid the provide evidence that the two active sAC proteins are generated a and are of distinct from of the truncated In we provide evidence that bicarbonate is a signal that regulates cAMP levels in to be this regulation an on and differentiation of these cells as well as on the of Cyclic AMP, the product of adenylyl cyclase, is an important second messenger controlling signaling in a wide variety of cells in almost all prokaryotic and eukaryotic organisms (1Sunahara R.K. Dessauer C.W. Gilman A.G. Annu. Rev. Pharmacol. Toxicol. 1996; 36: 461-480Crossref PubMed Scopus (732) Google Scholar, 2Taussig R. Gilman A.G. J. Biol. Chem. 1995; 270: 1-4Abstract Full Text Full Text PDF PubMed Scopus (431) Google Scholar). This also holds true for the male gonad, where cAMP signaling is essential for the hormonal regulation of somatic cell differentiation and function. Similarly, acquisition of motility, capacitation, and acrosome reaction, processes that render mature spermatozoa competent to fertilize an egg, are dependent on cAMP signaling (3Garbers D.L. Tubb D.J. Kopf G.S. Biol. Reprod. 1980; 22: 526-532Crossref PubMed Scopus (21) Google Scholar, 4Visconti P.E. Stewart-Savage J. Blasco A. Battaglia L. Miranda P. Kopf G.S. Tezon J.G. Biol. Reprod. 1999; 61: 76-84Crossref PubMed Scopus (183) Google Scholar, 5Yanagimachi R. Knobil E. Neill J. The Physiology of Reproduction. Raven Press, Ltd., New York1994: 189-317Google Scholar, 6Jaiswal B.S. Eisenbach M. Hardy D.M. Garbers D.L. Fertilization. Academic Press, San Diego2001Google Scholar). Conversely, little is known about the role of the cAMP pathway during the differentiation of germ cells. That cAMP signaling may be important for the spermatogenic cell is suggested by the genetic ablation of the CREB1-related protein CREM that is expressed at a high level in differentiating spermatids (7Delmas V. van der Hoorn F. Mellstrom B. Jegou B. Sassone-Corsi P. Mol. Endocrinol. 1993; 7: 1502-1514Crossref PubMed Scopus (187) Google Scholar). Mice deficient in CREM are sterile, and spermatogenesis is arrested at the beginning of spermatid differentiation (8Nantel F. Monaco L. Foulkes N.S. Masquilier D. LeMeur M. Henriksen K. A. M. Sassone-Corsi P. 1996; PubMed Scopus Google Scholar, E. 1996; PubMed Scopus Google Scholar). this and the that activated during differentiation cAMP-response in Sassone-Corsi P. Biol. Reprod. PubMed Scopus Google Scholar, Sassone-Corsi P. A. 1996; PubMed Scopus Google that CREM as a to the genetic spermatid differentiation P. Biol. PubMed Scopus Google Scholar). CREM is and activated by the protein in a to the cAMP-response element-binding protein (7Delmas V. van der Hoorn F. Mellstrom B. Jegou B. Sassone-Corsi P. Mol. Endocrinol. 1993; 7: 1502-1514Crossref PubMed Scopus (187) Google Scholar, J. J. Sassone-Corsi P. J. 1993; PubMed Scopus Google that cAMP signaling is for spermatogenesis. In male germ cells and mature cAMP is by at two a G adenylyl cyclase, is also expressed at high levels in the and in M. M. PubMed Google Scholar, M. Jegou B. J. PubMed Scopus Google Scholar). In known that a G and adenylyl cyclase is present in germ cells and spermatozoa A. PubMed Scopus Google Scholar, Cyclic Google Scholar, M. R. V. 1980; PubMed Scopus Google Scholar, K. V. J. PubMed Scopus (21) Google Scholar). The of the soluble form of adenylyl cyclase was by protein and J. L. A. 1999; PubMed Scopus Google Scholar). This adenylyl cyclase is an enzyme with and is to in J. L. A. 1999; PubMed Scopus Google Scholar). and not sAC activity J. L. A. 1999; PubMed Scopus Google a with the properties for the germ cell soluble cyclase M. R. V. 1980; PubMed Scopus Google Scholar, V. Mol. Endocrinol. PubMed Scopus Google Scholar, D. V. J. Endocrinol. PubMed Scopus Google Scholar). spermatozoa from all an adenylyl cyclase that is to bicarbonate A. J. Biol. Chem. Full Text PDF PubMed Google Scholar, P. J. 1993; PubMed Scopus Google Scholar). that sAC to the cyclase to bicarbonate in testis, in and in as a bicarbonate V. J. PubMed Scopus Google Scholar). of the cyclase identified a protein with a of V. Mol. Endocrinol. PubMed Scopus Google Scholar, D. V. J. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google Scholar). the of the sAC an open reading frame that for a protein J. L. A. 1999; PubMed Scopus Google Scholar). these that the protein is an that is to the active form J. L. A. 1999; PubMed Scopus Google Scholar). In of this of a truncated sAC the catalytic a active protein of J. L. A. 1999; PubMed Scopus Google Scholar). In the present we have this and provide evidence for an for the of the sAC we that two alternate transcripts are expressed in germ cells, and both the full-length and truncated to the germ cell adenylyl cyclase the high of bicarbonate in the seminiferous tubule in germ cells we that sAC is active on during germ cell differentiation and its bicarbonate regulation may provide a by germ cell is regulated by the evidence that two distinct transcripts encoding full-length and truncated as well as the corresponding proteins, are present in mouse and rat germ cells. expressed in a these proteins have distinct In we that the full-length sAC is active and to the of cAMP levels in germ cells and in mature the high of bicarbonate ions in the seminiferous tubule we that this cAMP not in mature spermatozoa also in differentiating of the properties of the soluble testis adenylyl cyclase that this activity in a of in rat testis V. Mol. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google in D. L. J. J. Biol. Chem. Full Text PDF PubMed Google and in testis D. V. J. Endocrinol. PubMed Scopus Google Scholar, L. V. PubMed Scopus Google Scholar). the of the sAC and the that the protein by this is that the form J. L. A. 1999; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). D. L. J. J. Biol. Chem. Full Text PDF PubMed Google have that of spermatozoa a soluble cyclase with a of properties of this product were not to of the to a soluble cyclase that is activated by bicarbonate have generated K. PubMed Scopus Google provide evidence that an may the a for sAC with an exon isolated during the of a rat testis J. L. A. 1999; PubMed Scopus Google by an with rat and mouse testis we have demonstrated that this is in a of of The high with these were that is not a a and RNase protection of RNA from of mouse and rat at different of that transcripts the for of the sAC that these transcripts are at be that of the sAC protein is is the truncated the that is active that of the adenylyl cyclase activity in the testis is to is to This on the of the two forms is with the on both the sAC activity from germ cells and the of testis. an that the full-length we that is not expressed in the testis of germ cells, and its is spermatogenesis to the spermatid provide evidence for an for the of full-length and truncated the that also the form as suggested by J. L. A. 1999; PubMed Scopus Google we that this for the presence of the two sAC of evidence that the is active in germ cells. cyclase activity was after of the full-length after immunoprecipitation with an that Both and are by the of chromatography of evidence for the presence of an active sAC form with a be that this of the presence of two distinct active is with V. Mol. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google Scholar, D. L. J. J. Biol. Chem. Full Text PDF PubMed Google Scholar). cAMP levels in cells were increased the was in as a with an of with little activity in the This with the distinct properties of the two the that the in the is generated by of The that as a the as an of two as a with an This to the in properties between the and the be that we the that and as and that immunoprecipitation with an that the also the active that the full-length sAC is and activated during the assay the presence of a is with the this provide evidence that both sAC are active in germ cells, the to be that as an for In to with the catalytic we that may be regulated by is that the of for and in the of These the full-length not the truncated protein by a in of the The in activity of the the is with this spermatozoa were as a of about of the soluble activity be that the full-length sAC is retained during the differentiation of spermatids and in a with the by V. J. PubMed Scopus Google Scholar). This the that both full-length and truncated sAC a role in the and of the generated that sAC is on during and that two active sAC proteins are present at in spermatids and This that sAC are not to the of cAMP levels in mature spermatozoa that sAC is active in cAMP during germ cell This is by the that bicarbonate cAMP levels in germ cells from 25- and 50-day-old rat testis, at a of mature spermatozoa are that seminiferous tubule a distinct from that of the present at levels in In the seminiferous tubule bicarbonate between 20 and J. Reprod. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. Endocrinol. PubMed Scopus Google Scholar, J. J. PubMed Scopus Google is that in the These are to sAC in a in germ cells as for cAMP In as activity in the testis, and in spermatozoa M. E. PubMed Scopus Google Scholar, K. PubMed Scopus Google Scholar, P. J. Mol. Reprod. 1999; PubMed Scopus Google Scholar, K. J. Biol. Reprod. 1999; 61: PubMed Scopus Google Scholar, E. L. PubMed Google Scholar). and rat germ cells transcripts P. J. Mol. Reprod. 1999; PubMed Scopus Google and high levels of activity have in cells of M. E. PubMed Scopus Google E. L. PubMed Google Scholar). all these and that bicarbonate cAMP in isolated germ cells, that bicarbonate cAMP during spermatid is not known bicarbonate during the seminiferous tubule we that bicarbonate in the seminiferous tubule is of an to cAMP for spermatid the provide evidence that the two active sAC proteins are generated a and are of distinct from of the truncated In we provide evidence that bicarbonate is a signal that regulates cAMP levels in to be this regulation an on and differentiation of these cells as well as on the of evidence that two distinct transcripts encoding full-length and truncated as well as the corresponding proteins, are present in mouse and rat germ cells. expressed in a these proteins have distinct In we that the full-length sAC is active and to the of cAMP levels in germ cells and in mature the high of bicarbonate ions in the seminiferous tubule we that this cAMP not in mature spermatozoa also in differentiating The of the properties of the soluble testis adenylyl cyclase that this activity in a of in rat testis V. Mol. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google in D. L. J. J. Biol. Chem. Full Text PDF PubMed Google and in testis D. V. J. Endocrinol. PubMed Scopus Google Scholar, L. V. PubMed Scopus Google Scholar). the of the sAC and the that the protein by this is that the form J. L. A. 1999; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). D. L. J. J. Biol. Chem. Full Text PDF PubMed Google have that of spermatozoa a soluble cyclase with a of properties of this product were not to of the to a soluble cyclase that is activated by bicarbonate have generated K. PubMed Scopus Google Scholar). provide evidence that an may the a for sAC with an exon isolated during the of a rat testis J. L. A. 1999; PubMed Scopus Google by an with rat and mouse testis we have demonstrated that this is in a of of The high with these were that is not a a and RNase protection of RNA from of mouse and rat at different of that transcripts the for of the sAC that these transcripts are at be that of the sAC protein is is the truncated the that is active that of the adenylyl cyclase activity in the testis is to is to This on the of the two forms is with the on both the sAC activity from germ cells and the of testis. an that the full-length we that is not expressed in the testis of germ cells, and its is spermatogenesis to the spermatid provide evidence for an for the of full-length and truncated the that also the form as suggested by J. L. A. 1999; PubMed Scopus Google we that this for the presence of the two sAC of evidence that the is active in germ cells. cyclase activity was after of the full-length after immunoprecipitation with an that Both and are by the of chromatography of evidence for the presence of an active sAC form with a be that this of the presence of two distinct active is with V. Mol. Endocrinol. PubMed Scopus Google Scholar, PubMed Scopus (21) Google Scholar, D. L. J. J. Biol. Chem. Full Text PDF PubMed Google Scholar). cAMP levels in cells were increased the was in cells. The as a with an of with little activity in the This with the distinct properties of the two the that the in the is generated by of The that as a the as an of two as a with an This to the in properties between the and the be that we the that and as and that immunoprecipitation with an that the also the active that the full-length sAC is and activated during the assay the presence of a is with the this provide evidence that both sAC are active in germ cells, the to be that as an for In to with the catalytic we that may be regulated by is that the of for and in the of These the full-length not the truncated protein by a in of the The in activity of the the is with this spermatozoa were as a of about of the soluble activity be that the full-length sAC is retained during the differentiation of spermatids and in a with the by V. J. PubMed Scopus Google Scholar). This the that both full-length and truncated sAC a role in the and of the generated that sAC is on during and that two active sAC proteins are present at in spermatids and This that sAC are not to the of cAMP levels in mature spermatozoa that sAC is active in cAMP during germ cell This is by the that bicarbonate cAMP levels in germ cells from 25- and 50-day-old rat testis, at a of mature spermatozoa are that seminiferous tubule a distinct from that of the present at levels in In the seminiferous tubule bicarbonate between 20 and J. Reprod. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. Endocrinol. PubMed Scopus Google Scholar, J. J. PubMed Scopus Google is that in the These are to sAC in a in germ cells as for cAMP In as activity in the testis, and in spermatozoa M. E. PubMed Scopus Google Scholar, K. PubMed Scopus Google Scholar, P. J. Mol. Reprod. 1999; PubMed Scopus Google Scholar, K. J. Biol. Reprod. 1999; 61: PubMed Scopus Google Scholar, E. L. PubMed Google Scholar). and rat germ cells transcripts P. J. Mol. Reprod. 1999; PubMed Scopus Google and high levels of activity have in cells of M. E. PubMed Scopus Google E. L. PubMed Google Scholar). all these and that bicarbonate cAMP in isolated germ cells, that bicarbonate cAMP during spermatid is not known bicarbonate during the seminiferous tubule we that bicarbonate in the seminiferous tubule is of an to cAMP for spermatid In the provide evidence that the two active sAC proteins are generated a and are of distinct from of the truncated In we provide evidence that bicarbonate is a signal that regulates cAMP levels in to be this regulation an on and differentiation of these cells as well as on the of for with the of for of sAC and R. for in also for in the of the