Maf1, repressor of tRNA transcription, is involved in the control of gluconeogenetic genes in Saccharomyces cerevisiae

Abstract

Maf1 is a negative regulator of RNA polymerase III (Pol III) in yeast. Maf1-depleted cells manifest elevated tRNA transcription and inability to grow on non-fermentable carbon source, such as glycerol. Using genomic microarray approach, we examined the effect of Maf1 deletion on expression of Pol II-transcribed genes in yeast grown in medium containing glycerol. We found that transcription of FBP1 and PCK1, two major genes controlling gluconeogenesis, was decreased in maf1Δ cells. FBP1 is located on chromosome XII in close proximity to a tRNA-Lys gene. Accordingly we hypothesized that decreased FBP1 mRNA level could be due to the effect of Maf1 on tgm silencing (tRNA gene mediated silencing). Two approaches were used to verify this hypothesis. First, we inactivated tRNA-Lys gene on chromosome XII by inserting a deletion cassette in a control wild type strain and in maf1Δ mutant. Second, we introduced a point mutation in the promoter of the tRNA-Lys gene cloned with the adjacent FBP1 in a plasmid and expressed in fbp1Δ or fbp1Δ maf1Δ cells. The levels of FBP1 mRNA were determined by RT-qPCR in each strain. Although the inactivation of the chromosomal tRNA-Lys gene increased expression of the neighboring FBP1, the mutation preventing transcription of the plasmid-born tRNA-Lys gene had no significant effect on FBP1 transcription. Taken together, those results do not support the concept of tgm silencing of FBP1. Other possible mechanisms are discussed.

Keywords: ATP; BSA; EDTA; FBP1; Gluconeogenesis; Maf1 repressor; ORF; PCK1; Pol II; Pol III; RNA polymerase II; RNA polymerase III; RT-qPCR; SDS; Yeast; adenosine tri-phosphate; bovine serum albumin; cDNA; complementary DNA; ethylenediaminetetraacetic acid; open reading frame; reverse transcription quantitative polymerase chain reaction; sodium dodecyl sulfate; tRNA transcription; tRNA-gene mediated silencing; tgm silencing; yeast gene encoding fructose-1,6-bisphosphatase; yeast gene encoding phosphoenolpyruvate carboxykinase.