Corradi, B. | bioRxiv Science

bioRxiv2025

REST/NRSF phosphorylation by CaMKIV regulates its transcriptional repressor activity and half-life

REST is a repressor of a large cluster of neural genes that homeostatically regulates neural activity. However, whether REST can be regulated by Ca2+ and Ca2+-activated kinases is unknown. We investigated Ca2+/calmodulin-dependent protein kinases (CaMKs) as upstream regulators of REST fate and activity. We show that REST is phosphorylated by CaMKIV at Ser-322 site located within the linker between the 5th-6th Zn-finger domains. Phosphomimic REST mutant in Serine-322 decreased REST repressor activity and caused its transition from nucleus to cytosol, followed by degradation. Molecular dynamics simulations of the phosphomimic N-terminal REST and the DNA RE1 sequence revealed a sharp decrease in the stability of the REST-RE1 binding interface. Moreover, the homeostatic effects of CaMKIV on the amplitude of excitatory synaptic currents were inhibited by the genetic deletion of REST. The results demonstrate that CaMKIV phosphorylation has a crucial role in the homeostatic regulation of REST levels and repressor activity. TeaserCaMKIV phosphorylation homeostatically regulates the repressor activity of REST on neural genes and its half-life.

neuroscience↗

bioRxiv2022

Molecular Dynamics Simulations of Ion Permeation in human NaV Channels

The recent determination of cryo-EM structures of voltage-gated sodium (Nav) channels has revealed many details of these proteins. However, knowledge of ionic permeation through the Nav pore remains limited. In this work, we performed atomistic molecular dynamics simulations to study the structural features of various neuronal Nav channels based on homology modeling of the cryo-EM structure of the human Nav1.4 channel and, in addition, on the more recent resolved configuration for Nav1.2. In particular, single Na+ permeation events during standard MD runs suggest that the ion resides in the inner part of the Nav selectivity filter (SF). On-the-fly free-energy parametrization (OTFP) temperature accelerated molecular dynamics (TAMD) was also used to calculate two-dimensional free energy surfaces (FESs) related to single/double Na+ translocation through the SF of the Nav1.2 homology model. The same thermodynamic analysis was performed on the cryo-EM based Nav1.2 configuration. These additional simulations revealed distinct mechanisms for single and double Na+ permeation through the wild-type SF, which has a charged lysine in the DEKA ring. In particular, the extracted protein-ion configurations are not accessible by the other modified SFs tested by TAMD/OTFP. Overall, the description of these mechanisms gives us new insights into ion conduction in human Nav cryo-EM based configurations, that could advance understanding of these systems and how they differ from potassium and bacterial Nav channels. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=181 SRC="FIGDIR/small/509656v1_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@53d852org.highwire.dtl.DTLVardef@9f17aaorg.highwire.dtl.DTLVardef@cacc7borg.highwire.dtl.DTLVardef@36bd64_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗