Cloning and characterization of the alternative promoter regions of the human LIMK2 gene responsible for alternative transcripts with tissue-specific expression.
We previously isolated the human LIMK2 gene and identified two alternative transcripts, LIMK2a and LIMK2b, which were differentially regulated in a tissue-specific manner. To examine this differential tissue-specific expression in detail, an RNase protection assay was performed, which demonstrated three expression patterns of the LIMK2 isoforms. In digestive organs, the LIMK2a transcripts were preferentially expressed in fetal and adult tissues; in brain and lung, the LIMK2a transcript was predominantly expressed only in fetal tissue, and in placenta, the LIMK2b transcript was expressed more abundantly than that of LIMK2a. To further investigate this mechanism and the transcription factors involved, we isolated the two distinct 5' upstream regions from the phage genomic library and found that both LIMK2a and 2b promoters have a single major transcription initiation site and the characteristics of a TATA-less promoter. A luciferase reporter assay of the transcriptional activity revealed positive as well as negative regulatory regions within both promoters. The co-transfection assay suggested that the MZF-1 might regulate the expression of the LIMK2 isoforms in a different manner. The RORalpha1 might also be involved in the transcriptional regulation of the LIMK2b isoform. The genomic structure of the LIMK2 gene was also determined. These findings should lead to a better understanding of the possibly diverse functions of the LIMK family.
Nomoto
S
S
Laboratory of Ultrastructure Research, Aichi Cancer Center Research Institute, 1-1 Kanokoden, Chikusa-ku, Nagoya 464-8681, Japan.
Tatematsu
Y
Y
Takahashi
T
T
Osada
H
H
D017398:Alternative Splicing; D001483:Base Sequence; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics*; D005786:Gene Expression Regulation; D015723:Gene Library; D017930:Genes, Reporter; D006801:Humans; D051741:Kruppel-Like Transcription Factors; D054461:Lim Kinases; D008957:Models, Genetic; D008969:Molecular Sequence Data; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1; D009928:Organ Specificity; D010920:Placenta / Q000378:metabolism; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms; D017346:Protein Serine-Threonine Kinases; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014018:Tissue Distribution; D015534:Trans-Activators / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D051741:Kruppel-Like Transcription Factors; C085969:MZF1 protein, human; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1; D020033:Protein Isoforms; D018160:Receptors, Cytoplasmic and Nuclear; D015534:Trans-Activators; D014157:Transcription Factors; C102895:LIMK2 protein, human; D054461:Lim Kinases; D017346:Protein Serine-Threonine Kinases