HSF1 and HSF2 in Stress and EMT-Associated Transcriptional Networks

Protein-damaging stress frequently occurs during the lifetime of a cell. To survive these insults, cells rely on the action of proteins called transcription factors that adjust the expression of genes, such as chaperones, according to the environmental requirement. Heat shock factors (HSFs) are key transcription factors that modulate gene expression upon proteotoxic insults; however, they are also important during several physiological processes related to development. To regulate transcription, HSFs undergo a complex activation process that involves nuclear accumulation, trimerization, and post-translational modifications (PTMs). However, there are big gaps in our knowledge regarding the importance of PTMs in the activation of HSFs, and it remains to be established if different HSF family members modulate transcription similarly during different stress conditions. Importantly, advances in our understanding of HSFs’ pro-survival roles have revealed that their misregulation can provoke pro-tumorigenic gene expression programs, which various cancers rely on to promote survival and invasion. This thesis comprises three studies that characterize different elements of HSF biology. The first study seeks to determine how phosphorylation in the regulatory domain of HSF1 affects its transcriptional capacity. The focus in the second study is placed on the ability of HSF1 and HSF2 to collaborate in the modulation of transcription, and it aims to map the network of genes and enhancers regulated by both HSF during heat and oxidative stress. The final study differs from the previous two in that it deals with the involvement of HSFs in the progression of cancer. Based on an earlier study showing that some cancer cells downregulate HSF2 to promote invasion, the third study aims primarily to uncover the relevant signaling pathway that suppresses the expression of HSF2. Also, the study seeks to determine why reduced levels of HSF2 are advantageous for certain cancer cells. Together, these studies characterize fundamental aspects of HSFs in stress regulation and explore how suppression of HSF2 may benefit cancer progression.Proteinskadande stress uppstår ofta under en cells livstid. För att överleva denna skada förlitar sig cellen på proteiner som kallas för transkriptionsfaktorer vilka justerar uttrycket av gener, inklusive chaperoner, i enlighet med miljöbehovet. Värmechockfaktorer (HSFs) är viktiga transkriptionsfaktorer som modulerar genuttryck vid proteinskadande stress, men de är också viktiga under flera fysiologiska processer relaterade till utveckling. För att reglera transkription genomgår HSF:s en komplex aktiveringsprocess som involverar nukleär ackumulering, trimerisering och post-translationella modifieringar (PTMs). Det finns dock stora luckor i vår kunskap om betydelsen av PTMs i aktiveringen av HSFs, och det återstår att fastställa om olika HSF-familjemedlemmar modulerar transkription på liknande sätt under olika stressförhållanden. I samband med framsteg i vår förståelse av HSFs roll i överlevnad har det framstått att deras felaktiga reglering kan stimulera uttrycket av flera gener som stöder utvecklingen av cancer, och olika typer av cancer förlitar sig på dessa nätverk av gener för att understöd deras överlevnad och invasionsförmåga. Denna avhandling sammanställer tre studier vars syfte är att karakterisera olika aspekter av HSF-biologi. Målet i den första studien är att fastställa hur fosforylering i den regulatoriska domänen av HSF1 påverkar dess transkriptionskapacitet. Fokuset i den andra studien är riktad på HSF1:s och HSF2:s förmåga att samarbeta i regleringen av transkription, och studien strävar till att kartlägga nätverket av gener och förstärkare, som regleras av både HSF under värme och oxidativ stress. Den sista studien skiljer sig från de två föregående i den meningen att den är inriktad på att utreda HSFs involvering i processer relaterat till cancer. På basen av en tidigare studie, som visade att vissa cancerceller minskar på uttrycket av HSF2 för att stimulera invasion, är det huvudsakliga målet i den tredje studien att utreda den relevanta signalräcka som kan styra uttrycket av HSF2. Dessutom strävar den tredje studien att bestämma varför låga nivåer av HSF2 är fördelaktiga för cancerceller. Sammantaget karakteriserar dessa studier grundläggande aspekter av HSFs i stressreglering och utforskar hur ett reducerat uttryck av HSF2 kan gynnar utvecklingen av cancer

HSFs drive transcription of distinct genes and enhancers during oxidative stress and heat shock

Reprogramming of transcription is critical for the survival under cellular stress. Heat shock has provided an excellent model to investigate nascent transcription in stressed cells, but the molecular mechanisms orchestrating RNA synthesis during other types of stress are unknown. We utilized PRO-seq and ChIP-seq to study how Heat Shock Factors, HSF1 and HSF2, coordinate transcription at genes and enhancers upon oxidative stress and heat shock. We show that pause-release of RNA polymerase II (Pol II) is a universal mechanism regulating gene transcription in stressed cells, while enhancers are activated at the level of Pol II recruitment. Moreover, besides functioning as conventional promoter-binding transcription factors, HSF1 and HSF2 bind to stress-induced enhancers to trigger Pol II pause-release from poised gene promoters. Importantly, HSFs act at distinct genes and enhancers in a stress type-specific manner. HSF1 binds to many chaperone genes upon oxidative and heat stress but activates them only in heat-shocked cells. Under oxidative stress, HSF1 localizes to a unique set of promoters and enhancers to trans-activate oxidative stress-specific genes. Taken together, we show that HSFs function as multi-stress-responsive factors that activate distinct genes and enhancers when encountering changes in temperature and redox state

CBP-HSF2 structural and functional interplay in Rubinstein-Taybi neurodevelopmental disorder

Rubinstein-Taybi syndrome (RSTS) is a neurodevelopmental disorder with unclear underlying mechanisms. Here, the authors unravel the contribution of a stress-responsive pathway to RSTS where impaired HSF2 acetylation, due to RSTS-associated CBP/EP300 mutations, alters the expression of neurodevelopmental players, in keeping with hallmarks of cell-cell adhesion defects.Patients carrying autosomal dominant mutations in the histone/lysine acetyl transferases CBP or EP300 develop a neurodevelopmental disorder: Rubinstein-Taybi syndrome (RSTS). The biological pathways underlying these neurodevelopmental defects remain elusive. Here, we unravel the contribution of a stress-responsive pathway to RSTS. We characterize the structural and functional interaction between CBP/EP300 and heat-shock factor 2 (HSF2), a tuner of brain cortical development and major player in prenatal stress responses in the neocortex: CBP/EP300 acetylates HSF2, leading to the stabilization of the HSF2 protein. Consequently, RSTS patient-derived primary cells show decreased levels of HSF2 and HSF2-dependent alteration in their repertoire of molecular chaperones and stress response. Moreover, we unravel a CBP/EP300-HSF2-N-cadherin cascade that is also active in neurodevelopmental contexts, and show that its deregulation disturbs neuroepithelial integrity in 2D and 3D organoid models of cerebral development, generated from RSTS patient-derived iPSC cells, providing a molecular reading key for this complex pathology.</p