Recent work has shown that DIG1
were derived from a single parental gene that existed prior to the whole-genome duplication (WGD) that occurred in the ancestor of S. cerevisiae
100-200 million years ago31
. Their continued presence in the genome suggests that their maintenance has an adaptive role. Indeed, previous work indicates that Dig1 and Dig2 inhibit Ste12 by interacting with distinct domains of the transcription factor, implying biochemical specialization 7,8
. However, their genetic redundancy for inhibiting Ste12 was puzzling. Studies presented here revealed three functions of Dig1 that are not redundant with those of Dig2: 1) control of gene expression noise, 2) regulation of the intranuclear distribution of Ste12, and 3) the control of long-range interactions between Ste12-target genes. We discuss below how these three functions may be related and the broader implications of these findings.
Dig1 is a well-studied regulatory protein that functions specifically in the pheromone response pathway and has only one reported biochemical function: to bind to a domain of Ste12 involved in protein-protein interactions5-8,32,33
. The loss of Dig1 is, therefore, expected solely to unshield protein-protein interaction domains on the Ste12 transcription factor. Although indirect mechanisms are always difficult to rule out, we propose that this unshielding induces aggregation of Ste12 molecules and target genes, which results in increased cell-to-cell variability in the basal output of the pheromone response pathway. Dig2, which binds the distinct DNA-binding domain of Ste127, 8
, does not share these functions. The aggregation of Ste12 molecules into one or two foci may create a domain within the nucleus where the transcription of Ste12-target genes can be activated. Our model suggests that the transcription of Ste12-target genes within the focus is more coordinated such that if one gene in the focus is transcribed, the others are, in turn, more likely to be expressed. Thus, such correlated expression within a single cell would be expected to yield increased correlated cell-to-cell variability in the transcriptional output of the pathway.
Transcriptional regulation that involves looping of DNA between distant sites via protein-protein interactions has been observed the lac
and λ phage39,40
. In the context of the results described here, it is notable that computational models of the lac
system suggest that gene regulation by DNA looping can affect fluctuations in transcription41
. These models predict that for transcriptional activators, DNA looping should increase noise in transcriptional outputs. Our model for the function of Dig1 is consistent with these theoretical predictions.
Recently, inter- and intrachromosomal interactions have been detected in other systems42-45
. In erythroid cells, for example, Klf1-regulated genes, including Hba
globin genes, display long-range inter- and intrachromosomal interactions42
. Although such interactions tend to correlate with transcriptional regulation and sites of active transcription, their precise functions remain a matter of considerable debate. Our observations suggest that while these long-range interactions could be important for gene expression, they may come at the cost of increased variability. This notion is in concordance with an emerging view that, in some cases, such gene interactions can be deleterious and even mutagenic46
. It will be interesting to explore whether mechanisms of noise regulation are pervasive among regulatory circuits that involve long-range DNA interactions and the extent to which gene localization is balanced with a need for limiting noise.
While establishing the generality of the effect of aggregate formation on output variability will require further investigation, we note that subcellular protein and DNA aggregation is not uncommon in biology. DNA replication and gene activation can occur in “factories” located at the nuclear periphery47-51
. Sites of DNA damage along with proteins that respond to DNA damage form nuclear foci in yeast52,53
. Telomeres are also known to cluster in the nucleus54
. Cytoplasmic P-bodies are foci of proteins involved in mRNA degradation and translational inhibition55-57
. Given our data, these foci may serve, in some cases, to promote simultaneity in cellular transactions. The development of assays that can distinguish between correlated and uncorrelated noise in these systems would allow the testing of such concepts.