cis elements that contribute to geminivirus transcriptional regulation and the efficiency of DNA replication

Abstract

The A genomic component of the geminivirus tomato golden mosaic virus (TGMV) contains a 5 intergenic sequence that includes the overlapping AL61 promoter and positive-strand origin of DNA replication. The TGMV AL1 protein negatively regulates its own transcription and mediates origin recognition by binding to a repeated motif shared by the AL61 promoter and the viral origin. We examined a series of truncated or mutated 5 intergenic regions in transient expression and replication assays to identify other DNA sequences that contribute to TGMV promoter and origin function. These experiments revealed that negative regulation of the AL61 promoter is complex, involving multiple cis-acting sequences and the AL1 and AL4 proteins, which acted through different DNA elements. We also found that mutation of the TATA box motif in the AL61 promoter reduced overall transcriptional activity and AL1-mediated repression, confirming the importance of this sequence in promoter function. Mutation of a G-box consensus sequence was highly detrimental to AL61 transcription and abolished AL1 sensitivity, suggesting that AL1 interferes with transcriptional activation. Cotransfection experiments showed that the TATA box and G-box motif mutations also impaired viral DNA replication in the presence of a wild-type origin but had no effect in its absence, demonstrating that these transcriptional motifs also function as replication efficiency elements.

AL1-dependent repression of transcription enhances expression of Tomato golden mosaic virus AL2 and AL3

“…DNA capable of expressing the AL1 protein with the amino acid alteration in AL4 (AL1 + /AL4 − ) had a small effect on repression of the AL62 promoter (data not shown). Our data is consistent with previous results demonstrating that AL4 has a relatively small effect on autoregulation of AL62 (Gröning et al, 1994;Eagle and Hanley-Bowdoin, 1997).This further demonstrates that AL4 repression plays a limited role in TGMV AL2 and AL3 gene expression.…”

Multi‐tasking of nonstructural gene products is required for bean yellow dwarf geminivirus transcriptional regulation

“…In the system described in this article, weak expression from the virion sense promoter was attributed to an absence of virus‐derived or virus‐activated cellular factors from the system. Previous experiments performed with WDV and MSV demonstrated that virion sense promoter activity is greater in phloem cells, suggesting that phloem‐specific transcription factors play a role in activating virion sense expression in the infection cycle [21–23]. In addition to this, a cell cycle specificity has been identified for both virion sense and complementary sense promoters of MSV [15].…”

“…The ability of Rep to down‐regulate expression of its own promoter has been studied previously. The AL1 protein of tomato golden mosaic virus, for example, has been shown to play a dual role in transcription and replication and it can inhibit its own expression by 20‐fold [4,22]. It is likely that either the modification of Rep, or the interaction of Rep with other viral or cellular proteins, may be involved in regulating the role of Rep as either a participant in viral replication or as a repressor of complementary sense gene expression.…”

“…The fact that Rep does not interact with RBR suggests that the C‐terminus of Rep hinders its ability to bind its LXCXE motif to the appropriate site in RBR. These steric differences between Rep and RepA may also explain how each have overlapping, but different, binding sites within the LIR of WDV [19,22]. While differential binding may also play a role in the inability of Rep to transactivate virion sense gene expression, the altered binding site of RepA may still have the same effects as Rep binding to inhibit complementary sense gene expression.…”

Molecular and biological characterization of Macroptilium yellow mosaic virus from Jamaica

“…A unique characteristic of the members in this clade is the G-Box motif, CACGTG, in their intergenic regions. The G-box is a cis-regulatory element of plant gene promoters and is also involved in the activation of the TGMV Rep gene (Eagle & Hanley-Bowdoin, 1997). In contrast, the DNA-A components of MaMPRV-[PR] and MaMPRV-[PR:Bean] occurring in Puerto Rico are more distantly related, despite being isolated from M. lathyroides and bean, respectively, and cluster with begomoviruses isolated from pepper and R. minima.…”