Bromodomain 4 (BRD4), a key regulator with pleiotropic functions, plays crucial roles in cancers and cellular stress responses. It exhibits dual functionality: chromatin-bound BRD4 regulates remodeling through its histone acetyltransferase (HAT) activity, while promoter-associated BRD4 regulates transcription through its kinase activity. Notably, chromatin-bound BRD4 lacks kinase activity, and RNA polymerase II (RNA Pol II)-bound BRD4 exhibits no HAT activity. This study unveils one mechanism underlying BRD4's functional switch. In response to diverse stimuli, c-Jun N-terminal kinase (JNK)-mediated phosphorylation of human BRD4 at Thr1186 and Thr1212 triggers its transient release from chromatin, disrupting its HAT activity and potentiating its kinase activity. Released BRD4 directly interacts with and phosphorylates RNA Pol II, PTEFb, and c-Myc, thereby promoting transcription of target genes involved in immune and inflammatory responses. JNK-mediated BRD4 functional switching induces CD8 expression in thymocytes and epithelial-to-mesenchymal transition (EMT) in prostate cancer cells. These findings elucidate the mechanism by which BRD4 transitions from a chromatin regulator to a transcriptional activator.
Published by Elsevier Inc.

Phosphorylation of cardiac myosin binding protein-C (cMyBP-C) is a determinant of cardiac myofilament function. Although cMyBP-C phosphorylation by various protein kinases has been extensively studied, the influence of protein phosphatases on cMyBP-C's multiple phosphorylation sites has remained largely obscure. Here we provide a detailed biochemical characterization of cMyBP-C dephosphorylation by protein phosphatases 1 and 2 A (PP1 and PP2A), and develop an integrated kinetic model for cMyBP-C phosphorylation using data for both PP1, PP2A and various protein kinases known to phosphorylate cMyBP-C. We find strong site-specificity and a hierarchical mechanism for both phosphatases, proceeding in the opposite direction of sequential phosphorylation by potein kinase A. The model is consistent with published data from human patients and predicts complex non-linear cMyBP-C phosphorylation patterns that are validated experimentally. Our results suggest non-redundant roles for PP1 and PP2A under both physiological and heart failure conditions, and emphasize the importance of phosphatases for cMyBP-C regulation.
© 2024. The Author(s).

The authors sought to understand sex differences in muscle metabolism in 73 older men and women.
Body composition, VO2max, and insulin sensitivity (M) by 3-hour hyperinsulinemic-euglycemic clamp with vastus lateralis muscle biopsies were measured.
Women had lower body weight, VO2max, and fat-free mass than men. Men had lower M, lower change (insulin minus basal) in muscle glycogen synthase (GS) activity, and lower change in AKT protein expression than women. M was associated with the change (insulin-basal) in GS activity and the change in AKT protein expression. Sex differences (n = 60) were tested with 6-month weight loss or 3×/week aerobic exercise training. The postintervention minus preintervention change (insulin-basal) (∆∆) in GS activity (fractional, independent, total) was higher in men than women in the weight loss group and ∆∆ in GS fractional activity was higher in women than men in the aerobic exercise group. In all participants, ∆∆ in GS fractional and independent activities was related to ∆∆ in AKT expression and glycogen content.
Sex differences in insulin sensitivity may be explained at the cellular muscle level, and to improve skeletal muscle insulin action in older adults, it may be necessary to recommend different behavioral strategies depending on the individual's sex.
Published 2024. This article is a U.S. Government work and is in the public domain in the USA.

Obscurin is a giant muscle protein (>800 kDa) featuring multiple signalling domains, including an SH3-DH-PH domain triplet from the Trio-subfamily of guanosine nucleotide exchange factors (GEFs). While previous research suggests that these domains can activate the small GTPases RhoA and RhoQ in cells, in vitro characterization of these interactions using biophysical techniques has been hampered by the intrinsic instability of obscurin GEF domains. To study substrate specificity, mechanism and regulation of obscurin GEF function by individual domains, we successfully optimized recombinant production of obscurin GEF domains and found that MST-family kinases phosphorylate the obscurin DH domain at Thr5798. Despite extensive testing of multiple GEF domain fragments, we did not detect any nucleotide exchange activity in vitro against 9 representative small GTPases. Bioinformatic analyses show that obscurin differs from other Trio-subfamily GEFs in several important aspects. While further research is necessary to evaluate obscurin GEF activity in vivo, our results indicate that obscurin has atypical GEF domains that, if catalytically active at all, are subject to complex regulation.
Copyright: © 2023 Koch et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Phosphorylation of cardiac myosin binding protein-C (cMyBP-C) is a crucial determinant of cardiac myofilament function. Although cMyBP-C phosphorylation by various protein kinases has been extensively studied, the influence of protein phosphatases on cMyBP-C’s multiple phosphorylation sites has remained largely obscure. Here we provide a detailed biochemical characterization of cMyBP-C dephosphorylation by protein phosphatases 1 and 2A (PP1 and PP2A) and develop an integrated kinetic model for cMyBP-C phosphorylation using data for both PP1, PP2A and protein kinases A (PKA), C and RSK2. We find strong site-specificity and a hierarchical mechanism for both phosphatases, proceeding in the opposite direction of sequential phosphorylation by PKA. The model is consistent with published data from human patients and predicts complex non-linear cMyBP-C phosphorylation patterns that are validated experimentally. Our results emphasize the importance of phosphatases for cMyBP-C regulation and prompt us to propose reciprocal relationships between cMyBP-C m-motif conformation, phosphorylation state and myofilament function.