Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal
Interest in extra-adrenal corticosteroid synthesis has been revived by technological advances and the quest for answers to clinical problems. The cytochrome P450 21-hydroxylase converts progesterone to deoxycorticosterone, the obligatory substrate for the production of the main adrenal steroids aldo...
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todo:paper_09600760_v62_n4_p277_Zhou2023-10-03T15:53:30Z Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal Zhou, M.-Y. Del Carmen Vila, M. Gomez-Sanchez, E.P. Gomez-Sanchez, C.E. complementary dna cytochrome p450 steroid 21 monooxygenase adrenal gland animal cell animal tissue aorta article brain brain stem cerebellum cho cell controlled study enzyme activity gene construct gene expression heart histochemistry hypothalamus kidney liver molecular cloning mutant nonhuman rat reverse transcription polymerase chain reaction sequence analysis sequence homology southern blotting Alternative Splicing Amino Acid Sequence Animals Base Sequence Cloning, Molecular COS Cells DNA, Complementary Molecular Sequence Data Rats Rats, Sprague-Dawley Sequence Alignment Sequence Homology, Amino Acid Sequence Homology, Nucleic Acid Steroid 21-Hydroxylase Steroids Zona Glomerulosa Animalia Interest in extra-adrenal corticosteroid synthesis has been revived by technological advances and the quest for answers to clinical problems. The cytochrome P450 21-hydroxylase converts progesterone to deoxycorticosterone, the obligatory substrate for the production of the main adrenal steroids aldosterone, cortisol and corticosterone. The rat P450 21-hydroxylase was cloned and two constructs, 21OH-5 and 21OH-6, sequenced. The constructs are similar, except that 21OH-6 has three additional major insertions of 64, 70 and 84 bp, a 3 bp deletion, and four extra base pairs immediately before the poly-A sequence. The entire coding region of 21OH-5 has 87 and 71% homology with the mouse and human 21-hydroxylase cDNA, respectively, whereas the encoded protein has 84 and 65% homology. Reverse transcriptase-polymerase chain reaction (RT-PCR) combined with Southern blot demonstrated expression of both transcripts in the kidney, aorta, liver, cerebellum, hypothalamus and brain stem, heart and cerebrum, but not the hippocampus, in addition to the adrenal, The entire coding region of 21OH-5 and the corresponding region of 21OH-6 including the three introns were cloned into pCR3 and the plasmids transiently transfected into COS-7 cells. Only 22OH-5 was translated into active protein, converting approximately 64% of 3H-progesterone to deoxycorticosterone in 2 h. Fil:Del Carmen Vila, M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. JOUR info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_09600760_v62_n4_p277_Zhou |
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Universidad de Buenos Aires |
| institution_str |
I-28 |
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R-134 |
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Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
complementary dna cytochrome p450 steroid 21 monooxygenase adrenal gland animal cell animal tissue aorta article brain brain stem cerebellum cho cell controlled study enzyme activity gene construct gene expression heart histochemistry hypothalamus kidney liver molecular cloning mutant nonhuman rat reverse transcription polymerase chain reaction sequence analysis sequence homology southern blotting Alternative Splicing Amino Acid Sequence Animals Base Sequence Cloning, Molecular COS Cells DNA, Complementary Molecular Sequence Data Rats Rats, Sprague-Dawley Sequence Alignment Sequence Homology, Amino Acid Sequence Homology, Nucleic Acid Steroid 21-Hydroxylase Steroids Zona Glomerulosa Animalia |
| spellingShingle |
complementary dna cytochrome p450 steroid 21 monooxygenase adrenal gland animal cell animal tissue aorta article brain brain stem cerebellum cho cell controlled study enzyme activity gene construct gene expression heart histochemistry hypothalamus kidney liver molecular cloning mutant nonhuman rat reverse transcription polymerase chain reaction sequence analysis sequence homology southern blotting Alternative Splicing Amino Acid Sequence Animals Base Sequence Cloning, Molecular COS Cells DNA, Complementary Molecular Sequence Data Rats Rats, Sprague-Dawley Sequence Alignment Sequence Homology, Amino Acid Sequence Homology, Nucleic Acid Steroid 21-Hydroxylase Steroids Zona Glomerulosa Animalia Zhou, M.-Y. Del Carmen Vila, M. Gomez-Sanchez, E.P. Gomez-Sanchez, C.E. Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal |
| topic_facet |
complementary dna cytochrome p450 steroid 21 monooxygenase adrenal gland animal cell animal tissue aorta article brain brain stem cerebellum cho cell controlled study enzyme activity gene construct gene expression heart histochemistry hypothalamus kidney liver molecular cloning mutant nonhuman rat reverse transcription polymerase chain reaction sequence analysis sequence homology southern blotting Alternative Splicing Amino Acid Sequence Animals Base Sequence Cloning, Molecular COS Cells DNA, Complementary Molecular Sequence Data Rats Rats, Sprague-Dawley Sequence Alignment Sequence Homology, Amino Acid Sequence Homology, Nucleic Acid Steroid 21-Hydroxylase Steroids Zona Glomerulosa Animalia |
| description |
Interest in extra-adrenal corticosteroid synthesis has been revived by technological advances and the quest for answers to clinical problems. The cytochrome P450 21-hydroxylase converts progesterone to deoxycorticosterone, the obligatory substrate for the production of the main adrenal steroids aldosterone, cortisol and corticosterone. The rat P450 21-hydroxylase was cloned and two constructs, 21OH-5 and 21OH-6, sequenced. The constructs are similar, except that 21OH-6 has three additional major insertions of 64, 70 and 84 bp, a 3 bp deletion, and four extra base pairs immediately before the poly-A sequence. The entire coding region of 21OH-5 has 87 and 71% homology with the mouse and human 21-hydroxylase cDNA, respectively, whereas the encoded protein has 84 and 65% homology. Reverse transcriptase-polymerase chain reaction (RT-PCR) combined with Southern blot demonstrated expression of both transcripts in the kidney, aorta, liver, cerebellum, hypothalamus and brain stem, heart and cerebrum, but not the hippocampus, in addition to the adrenal, The entire coding region of 21OH-5 and the corresponding region of 21OH-6 including the three introns were cloned into pCR3 and the plasmids transiently transfected into COS-7 cells. Only 22OH-5 was translated into active protein, converting approximately 64% of 3H-progesterone to deoxycorticosterone in 2 h. |
| format |
JOUR |
| author |
Zhou, M.-Y. Del Carmen Vila, M. Gomez-Sanchez, E.P. Gomez-Sanchez, C.E. |
| author_facet |
Zhou, M.-Y. Del Carmen Vila, M. Gomez-Sanchez, E.P. Gomez-Sanchez, C.E. |
| author_sort |
Zhou, M.-Y. |
| title |
Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal |
| title_short |
Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal |
| title_full |
Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal |
| title_fullStr |
Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal |
| title_full_unstemmed |
Cloning of two alternatively spliced 21-hydroxylase CDNAs from rat adrenal |
| title_sort |
cloning of two alternatively spliced 21-hydroxylase cdnas from rat adrenal |
| url |
http://hdl.handle.net/20.500.12110/paper_09600760_v62_n4_p277_Zhou |
| work_keys_str_mv |
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1807321171309363200 |