Hsf1 and Hsp70 constitute a two-component feedback loop that regulates the yeast heat shock response

Krakowiak, Joanna; Xu, Zheng; Patel, Nikit; Anandhakumar, Jayamani; Valerius, Kendra; Gross, David S.; Khalil, Ahmad S.; Pincus, David

doi:10.1101/183137

Cited by 26 publications

(50 citation statements)

References 39 publications

(29 reference statements)

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“…Our initial results are consistent with the longstanding view that misfolding of newly 405 synthesized polypeptides can serve as Hsf1 inducers (Baler, 1992;Masser et al, 2019), 406 presumably through recruitment of Hsp70 away from its repressive association with Hsf1 407 Krakowiak et al, 2018;Li et al, 2017). However, we have discovered an 408 alternative pathway for Hsf1 activation under conditions when newly synthesized proteins are 409 in short supply-when translational activity is low, such as following starvation or 410 pharmacological inhibition.…”

supporting

confidence: 80%

“…On the flip side, Hsf1 can be robustly activated 438 without a drop in pH, so long as cells are translationally active, indicating that acidification is 439 not necessary for Hsf1 activation. Recent key studies have demonstrated that production of 440 Hsp70 binding substrates that titrate Hsp70 away from Hsf1 suffices to induce Hsf1 in the 441 absence of heat shock Krakowiak et al, 2018). All these results are 442 consistent with the standard misfolding model: newly synthesized polypeptides misfold in 443 response to heat shock, leading to recruitment of Hsp70, which causes Hsf1 activation.…”

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confidence: 67%

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Transient intracellular acidification regulates the core transcriptional heat shock response

Triandafillou

Katanski

Dinner

et al. 2018

Preprint

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Heat shock induces a conserved transcriptional program regulated by heat shock factor 1 (Hsf1) in eukaryotic cells. Activation of this heat-shock response is triggered by heat-induced misfolding of newly synthesized polypeptides, and so has been thought to depend on ongoing protein synthesis. Here, using the the budding yeast Saccharomyces cerevisiae, we report the discovery that Hsf1 can be robustly activated when protein synthesis is inhibited, so long as cells undergo cytosolic acidification. Heat shock has long been known to cause transient intracellular acidification which, for reasons which have remained unclear, is associated with increased stress resistance in eukaryotes. We demonstrate that acidification is required for heat shock response induction in translationally inhibited cells, and specifically affects Hsf1 activation. Physiological heat-triggered acidification also increases population fitness and promotes cell cycle reentry following heat shock. Our results uncover a previously unknown adaptive dimension of the well-studied eukaryotic heat shock response. 1/45105 strain to express a red-fluorescent-protein-tagged version of Ssa4 (Ssa4-mCherry) from its 106 endogenous locus. Ssa4 is a strongly temperature-responsive Hsp70 chaperone, and its encoding 107 gene is a specific Hsf1 target (Hottiger et al., 1992;Pincus et al., 2018;Morano et al., 2012). 108This two-color reporter strain allowed us to simultaneously track intracellular pH and the stress 109 response at the single-cell level. 110 We stressed cells at 42 C for 20 minutes and then returned them to 30 C to recover. 111Samples were collected at 15-to 30-minute intervals during recovery and analyzed by flow 112 cytometry to monitor Ssa4-mCherry production. An example of the raw data, showing an 113 increase in fluorescence in the mCherry channel over time, is shown in Figure 1C. Although the 114 maturation time of the fluorophore, mCherry, confounds determination of the absolute timing 115 of the response, this delay is shared across experiments, allowing for direct comparison between 116 conditions and replicates. For each independent experiment, we tracked the median relative 117 change in red fluorescence over time, creating induction curves which characterize the response, 118as in Figure 1D. 119 5/45 6/45Intracellular acidification during heat shock promotes rapid heat-shock 120 protein production 121 With the tools in hand to quantify intracellular pH and induction of stress proteins, we set out 122 to first determine whether the ability to acidify during stress affected the cellular response. 123Evidence from the literature (Orij et al., 2011) strongly suggests that acidification results 124 primarily from influx of environmental protons, rather than (for example) the release of protons 125 from internal stores such as the vacuole. We confirmed a dependence on external protons by 126 heat-stressing cells in normal, acidic media (pH 4), or in media where the pH had been 127 adjusted to the cellular resting pH (7.5). Stressing cells i...

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supporting

confidence: 80%

mentioning

confidence: 67%

Transient intracellular acidification regulates the core transcriptional heat shock response

Triandafillou

Katanski

Dinner

et al. 2018

Preprint

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“…Previous models of the heat shock response at the cellular level aimed to characterize the importance and functionality of different aspects of the control circuit, including transcriptional, post-transcriptional and post-translational regulation, as well as the feedforward and feedback loops (Guisbert et al, 2008;Castells-Roca et al, 2011;Rieger et al, 2005;Petre et al, 2011;El-Samad et al, 2005;Abravaya et al, 1991;Krakowiak et al, 2017;Kurata et al, 2006). These models vary significantly in the level of detail at which different processes are modeled.…”

Section: Discussionmentioning

confidence: 99%

“…These models vary significantly in the level of detail at which different processes are modeled. In bacteria and yeast, the resolution of available experimental data allowed for assignment of functional roles to different branches of the regulatory circuits (El-Samad et al, 2005;Kurata et al, 2006) and the association of HSP-70 with HSF-1 (Krakowiak et al, 2017), respectively.…”

Section: Discussionmentioning

confidence: 99%

Dynamics of Heat Shock Detection and Response in the Intestine ofCaenorhabditis elegans

Levine

2019

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The heat shock response is the organized molecular response to stressors which disrupt proteostasis, potentially leading to protein misfolding and aggregation. While the regulation of the heat shock response is well-studied in single cells, its coordination at the cell, tissue, and systemic levels of a multicellular organism is poorly understood. To probe the interplay between systemic and cell-autonomous responses, we studied the upregulation of HSP-16.2, a molecular chaperone induced throughout the intestine of Caenorhabditis elegans following a heat shock, by taking longitudinal measurements in a microfluidic environment. Based on the dynamics of HSP-16.2 accumulation, we showed that a combination of heat shock temperature and duration define the intensity of stress inflicted on the worm and identified two regimes of low and high intensity stress. Modeling the underlying regulatory dynamics implicated the saturation of heat shock protein mRNA production in defining these two regimes and emphasized the importance of time separation between transcription and translation in establishing these dynamics. By applying a heat shock and measuring the response in separate parts of the animals, we implicated thermosensory neurons in accelerating the response and transducing information within the animal. We discuss possible implications of the systemic and cell level aspects and how they coordinate to facilitate the organismal response.

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“…Recently, multiple studies have pointed to Hsp70 as the primary and direct negative regulator of Hsf1 activity (15,16,22). Since Hsp70 is a major transcriptional target of Hsf1, these two proteins form a negative feedback loop that controls the dynamics of HSR induction (23). Although Hsp70 has emerged as a key repressor of Hsf1, roles for the other chaperones have not been ruled out.…”

Section: Introductionmentioning

confidence: 99%

Subcellular localization of the J-protein Sis1 regulates the heat shock response

Feder

Ali

Singh

et al. 2020

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One sentence summary: Localization of the J-protein Sis1 to a subcellular network of proteostasis factors activates the heat shock response. ABSTRACT Cells exposed to heat shock induce a conserved gene expression program -the heat shock response (HSR) -encoding chaperones like Hsp70 and other protein homeostasis (proteostasis) factors. Heat shock also triggers proteostasis factors to form subcellular quality control bodies, but the relationship between these spatial structures and the HSR is unclear.Here we show that localization of the J-protein Sis1 -a co-chaperone for Hsp70 -controls HSR activation in yeast. Under nonstress conditions, Sis1 is concentrated in the nucleoplasm where it promotes Hsp70 binding to the transcription factor Hsf1, repressing the HSR. Upon heat shock, Sis1 forms an interconnected network with other proteostasis factors that spans the nucleolus and the surface of the cortical ER. We propose that localization of Sis1 to this network directs Hsp70 activity away from Hsf1 in the nucleoplasm, leaving Hsf1 free to induce the HSR.In this manner, Sis1 couples HSR activation to the spatial organization of the proteostasis network. degradation (ERAD) (43). Together these images show that Sis1 colocalizes with RQC during heat shock.Beyond colocalization during heat shock, the 3D reconstructions of the lattice light sheet images revealed that Sis1-YFP, Hsp104-BFP, Rpn1-mScarlet, Ltn1-mScarlet and Cdc48-mScarlet form a semi-contiguous network ( Figure 5A-C). Sis1-YFP and Rpn1-mScarlet form an overlapping and highly ordered network that connects the nucleolar ring with a series of cytosolic foci ( Figure S4A, B, Movie S2). Hsp104 is excluded from the nucleolar ring but colocalizes with Sis1 and Rpn1 in the cytosolic network. Hsp90 (Hsc82-mScarlet) does not appear to participate in the network nor in any other Sis1-containing structures ( Figure S4C).The ER has been previously implicated as an organizational factor for the spatial arrangement of protein quality control factors, including Hsp104 (27,44). Moreover, we found that the ER structural protein Rtn1 co-precipitated with Sis1-3xFLAG during heat shock. To test if the Sis1 cytosolic network forms in proximity to the ER, we imaged Rtn1-mScarlet in cells expressing Sis1-YFP and Hsp104-BFP. Indeed, Sis1-YFP increased its signal overlap with Rtn1-mScarlet from MOC = 0.13 under nonstress conditions to MOC = 0.60 during heat shock ( Figure 5D), suggesting increased association with the ER. Space-filling 3D cell projections reveal the orientation of the interaction network, with Rtn1 toward the periphery and Sis1 and Hsp104 lining the interior ( Figure 5E). The imaging data suggest that Sis1 forms a highly connected network with the proteasome and RQC on the surface of the ER.Lastly, we tested if Sis1 is required for the proteasome to re-localize to the nucleolar ring during heat shock. To this end we anchored away Sis1 prior to heat shocking cells and monitored localization of Rpn1-mScarlet and Hsp104-BFP. Following Sis1 anchor away, we found that Rpn...

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Hsf1 and Hsp70 constitute a two-component feedback loop that regulates the yeast heat shock response

Abstract: 20Models for regulation of the eukaryotic heat shock response typically invoke a negative 21 feedback loop consisting of the transcriptional activator Hsf1 and a molecular chaperone 22

Cited by 26 publications

References 39 publications

Transient intracellular acidification regulates the core transcriptional heat shock response

Transient intracellular acidification regulates the core transcriptional heat shock response

Dynamics of Heat Shock Detection and Response in the Intestine ofCaenorhabditis elegans

Subcellular localization of the J-protein Sis1 regulates the heat shock response

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