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Down-regulated expression of atypical PKC-binding domain deleted asip isoforms in human hepatocellular carcinomas
1Laboratory of Molecular and Cellular Oncology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.
Abstract:
Asip is a mammalian homologue of polarity protein Par-3 of Caenorhabditis elegans and Bazooka of Drosophila melanogaster. Asip/Par-3/Bazooka are PDZ-motif containing proteins that localize asymmetrically to the cell periphery and play a pivotal role in cell polarity and asymmetric cell division. In the present study, we have cloned human asip cDNA and its splicing variants by 5'-RACE and RT-PCR using candidate human EST clones which have a high homology to rat asip cDNA. The full-length cDNA of human asip encodes a 1,353 aa protein exhibiting 88% similarity to the rat one. Human asip is a single copy gene consisting of at least 26 exons and localizing in human chromosome 10, band p11.2, with some extraordinarily long introns. All exon/intron boundary nucleotides conform to the "gt-ag" rule. Three main transcripts were detected by Northern blot analysis, and at least five variants, from alternative splicing and polyadenylation, have been identified by RT-PCR and liver cDNA library screening. Exon 17b deleted asip mRNAs expressed ubiquitously in normal human tissues, including liver, on RT-PCR analysis. However, they were absent from most human liver cancer cell lines examined. More interestingly, the expression of exon 17b deleted variants was down regulated in 52.6% (10/19) clinic specimens of human hepatocellular carcinomas (HCCs), compared with the surrounding nontumorous liver tissues from the same patients. The presence of various splicing transcripts, the variation of their distribution among different tissues and cells, and their differential expressions in human HCCs suggest that human Asip isoforms may function in different context.
Insights
Human ASIP (a cell polarity protein) has multiple splicing variants, with one variant notably absent in liver cancer cells and downregulated in hepatocellular carcinomas (HCCs). This suggests ASIP isoforms may play a role in HCC development.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- ASIP is a mammalian homolog of conserved cell polarity proteins (Par-3, Bazooka).
- These proteins are crucial for establishing cell polarity and asymmetric cell division.
- ASIP contains PDZ motifs and localizes to the cell periphery.
Purpose of the Study:
- To clone human ASIP cDNA and identify its splicing variants.
- To investigate the expression patterns of human ASIP isoforms.
- To determine the role of ASIP variants in human hepatocellular carcinoma (HCC).
Main Methods:
- Cloning of human ASIP cDNA and variants using 5'-RACE and RT-PCR.
- Northern blot analysis to detect main transcripts.
- RT-PCR and liver cDNA library screening to identify splicing variants.
- Analysis of ASIP expression in normal tissues and HCC specimens.
Main Results:
- Human ASIP encodes a 1,353 aa protein with 88% similarity to the rat homolog.
- Human ASIP gene is located on chromosome 10p11.2 and comprises at least 26 exons.
- At least five ASIP variants were identified through alternative splicing and polyadenylation.
- Exon 17b deleted ASIP mRNAs are ubiquitously expressed in normal tissues but absent in most HCC cell lines.
- Downregulation of exon 17b deleted variants was observed in 52.6% of HCC specimens compared to adjacent non-tumorous tissue.
Conclusions:
- Human ASIP exhibits significant alternative splicing, generating diverse isoforms.
- The differential expression of ASIP variants, particularly the exon 17b deleted form, in HCC suggests a potential role in tumorigenesis.
- ASIP isoforms may have context-dependent functions, potentially impacting cell polarity and cancer development.