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Temporal and spatial expression patterns of the hsp16 and ubq-1 genes in transgenic C. elegans

University of British Columbia

The expression of the small 16 kDa heat shock protein gene (hspl6 ) family and of the polyubiquitin encoding gene (ubq-1) in Caenorhabditis elegans has been examined by introducing lacZ fusions into the nematode by transformation. Transcription of the hspl6-lacZ transgenes was totally heat shock dependent and resulted in the rapid synthesis of detectable levels of β-gaIactosidase in most somatic tissues. Although the two hspl6 gene pairs of C. elegans are highly similar within both their coding and non-coding sequences, quantitative and qualitative differences in the spatial pattern of expression between gene pairs were observed. The hspl6-48 promoter was shown to direct greater expression of β-galactosidase in muscle and hypodermis while the hspl6-41 promoter was more efficient in intestine and pharyngeal tissue. Transgenes which eliminated one promoter from a gene pair were expressed- at reduced levels, particularly in post-embryonic stages, suggesting that the heat shock elements (HSEs) in the intergenic region of an hspl6 gene pair may act cooperatively to achieve high levels of expression of both genes. The hspl6 gene pairs are never constitutively expressed. In addition, their heat inducibility is developmentally restricted; they are not heat inducible during gametogenesis or early embryogenesis. The hspl6 genes represent the first fully inducible system in C. elegans to be characterized in detail at the molecular level, and the promoters of these genes should be useful for studying the action of tissue or developmentally regulated genes in this organism. Animals carrying a translational ubq-1 construct consisting of 938 bp of ubq-1 upstream sequences fused to lacZ (ubq93S-lacZ) expressed β-galactosidase in embryos and in a tissue general manner in 20% of staining L1 larvae. Somatic expression in later stages was usually confined to body muscle. Progressively larger deletions extending from the 5’ end of ubq938-lacZ did not significantly alter the pattern of expression until 827 bp of sequence had been removed. Thus sequences upstream of the transcriptional start site, including a G/C rich block and a sequence resembling a TATA box (GAATAA) are not required for ubq-1 expression. Moreover, a basal level of expression was maintained in embryos when 903 bp had been deleted. These results suggest that some of the regulatory elements required for efficient expression of ubq-1 may reside within the transcribed region of the gene; alternatively they must lie more than 1.7 kb upstream or 0.8 kb downstream of this region. PCR analysis indicates that RNA molecules transcribed from the ubq938-IacZ and ubq 827-lacZ transgenes are trans-spliced to SL1, as is ubq—1 RNA.

Thesis/Dissertation

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2008-12-20

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http://hdl.handle.net/2429/3273

Biochemistry and Molecular Biology

University of British Columbia

UBCV

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