Abstract
Nuclear actin has been implicated in dynamic chromatin rearrangements in diverse eukaryotes. In mammalian cells, it is required to reposition double-strand DNA breaks to enable homologous recombination repair and to enhance transcription by facilitating RNA Pol II recruitment to gene promoters. In the yeast Saccharomyces cerevisiae, nuclear actin modulates interphase chromosome dynamics and is required to reposition the induced INO1 gene to the nuclear periphery. Here, we have investigated the role of actin in driving intergenic interactions between Heat Shock Factor 1 (Hsf1)-regulated Heat Shock Protein (HSP) genes in budding yeast. These genes, dispersed on multiple chromosomes, dramatically reposition following exposure of cells to acute thermal stress, leading to their clustering within dynamic biomolecular condensates. Using an auxin-induced degradation strategy, we found that conditional depletion of nucleators of either linear or branched F-actin (Bni1/Bnr1 and Arp2, respectively) had little or no effect on heat shock-induced HSP gene coalescence or transcription. In addition, we found that pretreatment of cells with latrunculin A, an inhibitor of both filamentous and monomeric actin, failed to affect intergenic interactions between activated HSP genes and their heat shock-induced intragenic looping and folding. Moreover, latrunculin A pretreatment had little effect on HSP gene expression at either RNA or protein levels. In notable contrast, we confirmed that repositioning of activated INO1 to the nuclear periphery and its proper expression do require actin. Collectively, our work suggests that transcriptional activation and 3D genome restructuring of thermally induced, Hsf1-regulated genes can occur in the absence of actin.