The copy numbers of many plant transcription factor (TF) genes substantially increased during terrestrialization. This allowed TFs to acquire new specificities and thus create gene regulatory networks (GRNs) with new biological functions to help plants adapt to terrestrial environments. Through characterizing Heat Shock Factor (HSF) genes MpHSFA1 and MpHSFB1 in the liverwort Marchantia polymorpha, we explored how heat-responsive GRNs widened their functions in M. polymorpha and Arabidopsis thaliana. An interspecies comparison of heat-induced transcriptomes and the evolutionary rates of HSFs demonstrated the emergence and subsequent rapid evolution of HSFB prior to terrestrialization. Transcriptome and metabolome analyses of M. polymorpha HSF-null mutants revealed that MpHSFA1 controls canonical heat responses such as thermotolerance and metabolic changes. MpHSFB1 also plays essential roles in heat responses, as well as regulating developmental processes including meristem branching and antheridiophore formation. Analysis of cis-regulatory elements revealed development- and stress-related TFs that function directly or indirectly downstream of HSFB. Male gametophytes of M. polymorpha showed higher levels of thermotolerance than female gametophytes, which could be explained by different expression levels of MpHSFA1U and MpHSFA1V on sex chromosome. We propose that the diversification of HSFs is linked to the expansion of HS responses, which enabled coordinated multicellular reactions in land plants.

VAMP-associated proteins (VAPs) are highly conserved endoplasmic reticulum (ER) resident proteins that establish ER contacts with multiple membrane compartments in many eukaryotes. However, VAP-mediated membrane tethering mechanisms remain ambiguous. Here, focusing on fission yeast ER-plasma membrane (PM) contact formation, using systematic interactome analyses and quantitative microscopy, we predict a non-protein-protein binding-based tethering mechanism of VAPs. We further demonstrate that VAP-anionic phospholipids interactions underlie ER-PM association and define the pH-responsive nature of VAP-tethered membrane contacts. Importantly, such conserved interactions, particularly with phosphatidylinositol 4-phosphate (PI4P) and phosphatidylinositol (PI), are defective in amyotrophic lateral sclerosis (ALS)-associated human VAPB mutant. Moreover, we identify a conserved FFAT-like motif locating at the autoinhibitory hotspot of the essential PM proton pump Pma1. This modulatory VAP-Pma1 interaction is crucial for pH homeostasis. We thus propose an ingenious strategy for maintaining intracellular pH by coupling Pma1 modulation with pH-biosensory ER-PM contacts via VAP-mediated interactions.