Expression of human pyruvate carboxylase in insect cells using the baculovirus system

Natthida Petchampai, Vichai Boonsaeng, John C. Wallace and Sarawut Jitrapakde

Structural and promoter regions of the murine pyruvate carboxylase gene

We have cloned and sequenced the gene encoding mouse pyruvate carboxylase (mPC) [EC 6.4.1.1]. The coding region contains 19 exons, one 5′-untranslated region exon, and 19 introns in 22 kb of genomic DNA. This gene's exon/intron organization is highly conserved with respect to rat and human PC genes. The mPC gene promoter lacks canonical TATA and CCAAT boxes, in common with a number of housekeeping genes. Transient expressions in COS-1 of a luciferase reporter gene under the control of 5′-nested deletions of the 5′-flanking sequence of the mPC gene have identified the 166-bp minimal sequence required for basal transcription. Alternative splicing at the 5′-untranslated region exon of the mouse PC gene results in the production of two alternate transcripts bearing different 5′-noncoding regions. Both transcripts are highly expressed in kidney and liver and moderately expressed in heart and testis and expressed at a low level in spleen.Sarawut Jitrapakdee; Nathida Petchamphai; Piyanate Sunyakumthorn; John C. Wallace; Vichai Boonsaen