Characterization of the human CALM2 calmodulin gene and comparison of the transcriptional activity of CALM1, CALM2 and CALM3.
Human calmodulin is encoded by three genes CALM1, CALM2 and CALM3 located on different chromosomes. To complete the characterization of this family, the exon-intron structure of CALM2 was solved by a combination of genomic DNA library screening and genomic PCR amplification. Intron interruptions were found at identical positions in human CALM2 as in CALM1 and CALM3; however, the overall size of CALM2 (16 kb) was almost twice that of the other two human CALM genes. Over 1 kb of the 5' flanking sequence of human CALM2 were determined, revealing the presence of a TATA-like sequence 27 nucleotides upstream of the transcriptional start site and several conserved sequence elements possibly involved in the regulation of this gene. To determine if differential transcriptional activity plays a major role in regulating cellular calmodulin levels, we directly measured and compared the mRNA abundance and transcriptional activity of the three CALM genes in proliferating human teratoma cells. CALM3 was at least 5-fold more actively transcribed than CALM1 or CALM2. CALM transcriptional activity agreed well with the mRNA abundance profile in the teratoma cells. In transient transfections using luciferase reporter genes driven by 1 kb of the 5' flanking DNA of the three CALM genes, the promoter activity correlated with the endogenous CALM transcriptional activity, but only when the 5' untranslated regions were included in the constructs. We conclude that the CALM gene family is differentially active at the transcriptional level in teratoma cells and that the 5' untranslated regions are necessary to recover full promoter activation.
Toutenhoofd
S L
SL
Department of Biochemistry and Molecular Biology, Mayo Graduate School, Mayo Foundation, Rochester, MN 55905, USA.
Foletti
D
D
Wicki
R
R
Rhyner
J A
JA
Garcia
F
F
Tolon
R
R
Strehler
E E
EE
D000595:Amino Acid Sequence; D001483:Base Sequence; D002147:Calmodulin / Q000235:genetics*; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005091:Exons / Q000235:genetics; D005786:Gene Expression Regulation; D015723:Gene Library; D005796:Genes; D017930:Genes, Reporter; D006801:Humans; D007438:Introns / Q000235:genetics; D008969:Molecular Sequence Data; D005810:Multigene Family; D009928:Organ Specificity; D016133:Polymerase Chain Reaction; D012326:RNA Splicing; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D018243:Teratocarcinoma / Q000473:pathology; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D002147:Calmodulin; D018076:DNA, Complementary; D012333:RNA, Messenger; D015500:Chloramphenicol O-Acetyltransferase