The asymmetric distribution of RNA polymerase II and nucleosomes on replicated daughter genomes is caused by differences in replication timing between the lagging and the leading strand.

Chromatin features are thought to have a role in the epigenetic transmission of transcription states from one cell generation to the next. It is unclear how chromatin structure survives disruptions caused by genomic replication or whether chromatin features are instructive of the transcription state of the underlying gene. We developed a method to monitor budding yeast replication, transcription, and chromatin maturation dynamics on each daughter genome in parallel, with which we identified clusters of secondary origins surrounding known origins. We found a difference in the timing of lagging and leading strand replication on the order of minutes at most yeast genes. We propose a model in which the majority of old histones and RNA polymerase II (RNAPII) bind to the gene copy that replicated first, while newly synthesized nucleosomes are assembled on the copy that replicated second. RNAPII enrichment then shifts to the sister copy that replicated second. The order of replication is largely determined by genic orientation: If transcription and replication are codirectional, the leading strand replicates first; if they are counterdirectional, the lagging strand replicates first. A mutation in the Mcm2 subunit of the replicative helicase Mcm2-7 that impairs Mcm2 interactions with histone H3 slows down replication forks but does not qualitatively change the asymmetry in nucleosome distribution observed in the WT. We propose that active transcription states are inherited simultaneously and independently of their underlying chromatin states through the recycling of the transcription machinery and old histones, respectively. Transcription thus actively contributes to the reestablishment of the active chromatin state.

High prevalence of malnutrition in systemic sclerosis: Results from a French monocentric cross-sectional study.

OBJECTIVES: Systemic sclerosis (SSc) can cause malnutrition due to frequent gastrointestinal involvement. However, prevalence of malnutrition in SSc is poorly known. The aim of this study was to evaluate the prevalence of malnutrition in SSc and its potential associations with disease features in patients from a tertiary referral center. METHODS: All patients meeting American College of Rheumatology/European Alliance of Associations for Rheumatology criteria for SSc followed between January 1, 1985, and January 1, 2019, at the Department of Internal Medicine, Saint Eloi University Hospital, were included. Malnutrition was assessed using the 2020 French recommendations for SSc and the malnutrition universal screening tool score. Severe malnutrition was defined via the French Haute Autorité de Santé (National Health Authority) 2007 criteria. RESULTS: A total of 120 patients were included, with mean age 64 (± 15) y and a female-to-male sex ratio of 5:1. According to 2020 French recommendations, 71 patients (59.2%) were malnourished and 30 (25%) had at least one criterion of severe malnutrition. With the malnutrition universal screening tool score, 41.7%, 20%, and 38.3%, respectively, had low, medium, and high risk of malnutrition. Multivariate analysis revealed the following results: 1) malnutrition was associated with cardiac involvement (P < 0.01); 2) a high malnutrition universal screening tool score was also associated with specific cardiac involvement (P < 0.01); and 3) severe malnutrition was strongly correlated with interincisal distance <35 mm (P = 0.02). CONCLUSIONS: Malnutrition affects more than half of SSc patients and is associated with specific cardiac involvement. Interincisal distance <35 mm could be a red flag for severe malnutrition in SSc.

Cell membrane expression of cardiac sodium channel Na(v)1.5 is modulated by alpha-actinin-2 interaction.

Cardiac sodium channel Na(v)1.5 plays a critical role in heart excitability and conduction. The molecular mechanism that underlies the expression of Na(v)1.5 at the cell membrane is poorly understood. Previous studies demonstrated that cytoskeleton proteins can be involved in the regulation of cell surface expression and localization of several ion channels. We performed a yeast two-hybrid screen to identify Na(v)1.5-associated proteins that may be involved in channel function and expression. We identified alpha-actinin-2 as an interacting partner of the cytoplasmic loop connecting domains III and IV of Na(v)1.5 (Na(v)1.5/LIII-IV). Co-immunoprecipitation and His(6) pull-down assays confirmed the physical association between Na(v)1.5 and alpha-actinin-2 and showed that the spectrin-like repeat domain is essential for binding of alpha-actinin-2 to Na(v)1.5. Patch-clamp studies revealed that the interaction with alpha-actinin-2 increases sodium channel density without changing their gating properties. Consistent with these findings, coexpression of alpha-actinin-2 and Na(v)1.5 in tsA201 cells led to an increase in the level of expression of Na(v)1.5 at the cell membrane as determined by cell surface biotinylation. Lastly, immunostaining experiments showed that alpha-actinin-2 was colocalized with Na(v)1.5 along the Z-lines and in the plasma membrane. Our data suggest that alpha-actinin-2, which is known to regulate the functional expression of the potassium channels, may play a role in anchoring Na(v)1.5 to the membrane by connecting the channel to the actin cytoskeleton network.

Regulation of Na v channels in sensory neurons.

Voltage-gated Na(+) channels have an essential role in the biophysical properties of nociceptive neurons. Factors that regulate Na(+) channel function are of interest from both pathophysiological and therapeutic perspectives. Increasing evidence indicates that changes in expression or inappropriate modulation of these channels leads to electrical instability of the cell membrane and the inappropriate spontaneous activity that is observed following nerve injury, and that this might contribute to neuropathic pain. The role of Na(v) channels in nociception depends on modulation by factors such as auxiliary beta-subunits, cytoskeletal proteins and the phosphorylation state of neurons. In this review we describe the modulation of Na(v) channels on sensory neurons by auxiliary beta-subunits, protein kinases and cytoskeletal proteins.

Dynamics of Nucleosome Positioning Maturation following Genomic Replication.

Chromatin is thought to carry epigenetic information from one generation to the next, although it is unclear how such information survives the disruptions of nucleosomal architecture occurring during genomic replication. Here, we measure a key aspect of chromatin structure dynamics during replication-how rapidly nucleosome positions are established on the newly replicated daughter genomes. By isolating newly synthesized DNA marked with 5-ethynyl-2'-deoxyuridine (EdU), we characterize nucleosome positions on both daughter genomes of S. cerevisiae during chromatin maturation. We find that nucleosomes rapidly adopt their mid-log positions at highly transcribed genes, which is consistent with a role for transcription in positioning nucleosomes in vivo. Additionally, experiments in hir1Δ mutants reveal a role for HIR in nucleosome spacing. We also characterized nucleosome positions on the leading and lagging strands, uncovering differences in chromatin maturation dynamics at hundreds of genes. Our data define the maturation dynamics of newly replicated chromatin and support a role for transcription in sculpting the chromatin template.

Lidocaine promotes the trafficking and functional expression of Na(v)1.8 sodium channels in mammalian cells.

Nociceptive neurons of the dorsal root ganglion (DRG) express a combination of rapidly gating TTX-sensitive and slowly gating TTX-resistant Na currents, and the channels that produce these currents have been cloned. The Na(v)1.7 and Na(v)1.8 channels encode for the rapidly inactivating TTX-sensitive and slowly inactivating TTX-resistant Na currents, respectively. Although the Na(v)1.7 channel expresses well in cultured mammalian cell lines, attempts to express the Na(v)1.8 channel using similar approaches has been met with limited success. The inability to heterologously express Na(v)1.8 has hampered detailed characterization of the biophysical properties and pharmacology of these channels. In this study, we investigated the determinants of Na(v)1.8 expression in tsA201 cells, a transformed variant of HEK293 cells, using a combination of biochemistry, immunochemistry, and electrophysiology. Our data indicate that the unusually low expression levels of Na(v)1.8 in tsA201 cells results from a trafficking defect that traps the channel protein in the endoplasmic reticulum. Incubating the cultured cells with the local anesthetic lidocaine dramatically enhanced the cell surface expression of functional Na(v)1.8 channels. The biophysical properties of the heterologously expressed Na(v)1.8 channel are similar but not identical to those of the TTX-resistant Na current of native DRG neurons, recorded under similar conditions. Our data indicate that the lidocaine acts as a molecular chaperone that promotes efficient trafficking and increased cell surface expression of Na(v)1.8 channels.