Nuclear F-actin assembly on damaged chromatin is regulated by DYRK1A and Spir1 phosphorylation.

Nuclear actin-based movements support DNA double-strand break (DSB) repair. However, molecular determinants that promote filamentous actin (F-actin) formation on the damaged chromatin remain undefined. Here we describe the DYRK1A kinase as a nuclear activity that promotes local F-actin assembly to support DSB mobility and repair, accomplished in part by its targeting of actin nucleator spire homolog 1 (Spir1). Indeed, perturbing DYRK1A-dependent phosphorylation of S482 mis-regulated Spir1 accumulation at damaged-modified chromatin, and led to compromised DSB-associated actin polymerization and attenuated DNA repair. Our findings uncover a role of the DYRK1A-Spir1 axis in nuclear actin dynamics during early DSB responses, and highlight the intricate details of nuclear cytoskeletal network in DSB repair and genome stability maintenance.

Microtubule-Mediated Regulation of ß2AR Translation and Function in Failing Hearts.

BACKGROUND: ß1AR (beta-1 adrenergic receptor) and ß2AR (beta-2 adrenergic receptor)-mediated cyclic adenosine monophosphate signaling has distinct effects on cardiac function and heart failure progression. However, the mechanism regulating spatial localization and functional compartmentation of cardiac ß-ARs remains elusive. Emerging evidence suggests that microtubule-dependent trafficking of mRNP (messenger ribonucleoprotein) and localized protein translation modulates protein compartmentation in cardiomyocytes. We hypothesized that ß-AR compartmentation in cardiomyocytes is accomplished by selective trafficking of its mRNAs and localized translation. METHODS: The localization pattern of ß-AR mRNA was investigated using single molecule fluorescence in situ hybridization and subcellular nanobiopsy in rat cardiomyocytes. The role of microtubule on ß-AR mRNA localization was studied using vinblastine, and its effect on receptor localization and function was evaluated with immunofluorescent and high-throughput Förster resonance energy transfer microscopy. An mRNA protein co-detection assay identified plausible ß-AR translation sites in cardiomyocytes. The mechanism by which ß-AR mRNA is redistributed post-heart failure was elucidated by single molecule fluorescence in situ hybridization, nanobiopsy, and high-throughput Förster resonance energy transfer microscopy on 16 weeks post-myocardial infarction and detubulated cardiomyocytes. RESULTS: ß1AR and ß2AR mRNAs show differential localization in cardiomyocytes, with ß1AR found in the perinuclear region and ß2AR showing diffuse distribution throughout the cell. Disruption of microtubules induces a shift of ß2AR transcripts toward the perinuclear region. The close proximity between ß2AR transcripts and translated proteins suggests that the translation process occurs in specialized, precisely defined cellular compartments. Redistribution of ß2AR transcripts is microtubule-dependent, as microtubule depolymerization markedly reduces the number of functional receptors on the membrane. In failing hearts, both ß1AR and ß2AR mRNAs are redistributed toward the cell periphery, similar to what is seen in cardiomyocytes undergoing drug-induced detubulation. This suggests that t-tubule remodeling contributes to ß-AR mRNA redistribution and impaired ß2AR function in failing hearts. CONCLUSIONS: Asymmetrical microtubule-dependent trafficking dictates differential ß1AR and ß2AR localization in healthy cardiomyocyte microtubules, underlying the distinctive compartmentation of the 2 ß-ARs on the plasma membrane. The localization pattern is altered post-myocardial infarction, resulting from transverse tubule remodeling, leading to distorted ß2AR-mediated cyclic adenosine monophosphate signaling.

Multi-target regulatory mechanism of Yang Xin Tang - a traditional Chinese medicine against dementia.

BACKGROUND: Yang Xin Tang (YXT) is a traditional Chinese herbal preparation which has been reported to improve cognitive function and memory in patients with dementia. As the underlying mechanism of action of YXT has not been elucidated, we examined the effects of YXT and its major herbal components in regulating gene transcription and molecular targets related to Alzheimer's disease (AD). METHODS: Aqueous and ethanol extracts of YXT and selected herbal components were prepared and validated by standard methods. A series of biochemical and cellular assays were employed to assess the ability of the herbal extracts to inhibit acetylcholinesterase, reduce ß-amyloid aggregation, stimulate the differentiation of neural progenitor cells, suppress cyclooxygenase, and protect neurons against ß-amyloid or N-methyl-D-aspartate-induced cytotoxicity. The effects of YXT on multiple molecular targets were further corroborated by a panel of nine reporter gene assays. RESULTS: Extracts of YXT and two of its constituent herbs, Poria cocos and Poria Sclerotium pararadicis, significantly inhibited ß-amyloid aggregation and ß-amyloid-induced cytotoxicity. A protective effect of the YXT extract was similarly observed against N-methyl-D-aspartate-induced cytotoxicity in primary neurons, and this activity was shared by extracts of Radix Astragali and Rhizoma Chuanxiong. Although the YXT extract was ineffective, extracts of Poria cocos, Poria Sclerotium pararadicis and Radix Polygalae inhibited acetylcholine esterase, with the latter also capable of upregulating choline acetyltransferase. YXT and its components significantly inhibited the activities of the pro-inflammatory cyclooxygenases. Additionally, extracts of YXT and several of its constituent herbs significantly stimulated the phosphorylation of extracellular signal-regulated kinases and cAMP-responsive element binding protein, two molecular targets involved in learning and memory, as well as in the regulation of neurogenesis. CONCLUSIONS: Several constituents of YXT possess multiple regulatory effects on known therapeutic targets of AD that range from ß-amyloid to acetylcholinesterase. The demonstrated neuroprotective and neurogenic actions of YXT lend credence to its use as an alternative medicine for treating AD.

Latent membrane protein 1 and macrophage-derived TNFα synergistically activate and mobilize invadopodia to drive invasion of nasopharyngeal carcinoma.

Invadopodia are actin-rich membrane protrusions that digest the matrix barrier during cancer metastasis. Since the discovery of invadopodia, they have been visualized as localized and dot-like structures in different types of cancer cells on top of a 2D matrix. In this investigation of Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC), a highly invasive cancer frequently accompanied by neck lymph node and distal organ metastases, we revealed a new form of invadopodium with mobilizing features. Integration of live-cell imaging and molecular assays revealed the interaction of macrophage-released TNFα and EBV-encoded latent membrane protein 1 (LMP1) in co-activating the EGFR/Src/ERK/cortactin and Cdc42/N-WASP signaling axes for mobilizing the invadopodia with lateral movements. This phenomenon endows the invadopodia with massive degradative power, visualized as a shift of focal dot-like digestion patterns on a 2D gelatin to a dendrite-like digestion pattern. Notably, single stimulation of either LMP1 or TNFα could only enhance the number of ordinary dot-like invadopodia, suggesting that the EBV infection sensitizes the NPC cells to form mobilizing invadopodia when encountering a TNFα-rich tumor microenvironment. This study unveils the interplay of EBV and stromal components in driving the invasive potential of NPC via unleashing the propulsion of invadopodia in overcoming matrix hurdles. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Centrosomal protein TAX1BP2 inhibits centrosome-microtubules aberrations induced by hepatitis B virus X oncoprotein.

Liver cancer (hepatocellular carcinoma, HCC) is one of the most prevalent cancers worldwide. Several etiological factors of HCC, including hepatitis B or hepatitis C virus infection, liver cirrhosis and aflatoxin B1 intake has been identified. HBx, which is an oncogenic protein encoded by the hepatitis B virus, is strongly associated with hepatocarcinogenesis. Using stable HBx-expressing cell, we showed that HBx induced chromosome gain, with amplification of centrosomes numbers and deregulation of centrosome ultrastructure. To dissect the mechanism for chromosome instability, our result revealed that HBx contributed to a hyperactive centrosome-microtubule dynamics by accelerating microtubule nucleation and polymerization. Further investigations suggested that HBx interacted with a centrosome linker protein TAX1BP2, which has previously been shown to function as an intrinsic block of centrosome amplification and a tumour suppressor in HCC. Restoring TAX1BP2 was able to block HBx-mediated centrosome amplification and abolish the HBx-mediated centrosome aberration, thereby suppressing chromosome instability. Thus, we demonstrate here a mechanism by which HBx deregulates centrosome-microtubule dynamics through interacting with TAX1BP2, which underlines the possibility of restoration of TAX1BP2 to rescue cells from chromosome instability.

Regulator of G protein signaling 20 enhances cancer cell aggregation, migration, invasion and adhesion.

Several RGS (regulator of G protein signaling) proteins are known to be upregulated in a variety of tumors but their roles in modulating tumorigenesis remain undefined. Since the expression of RGS20 is elevated in metastatic melanoma and breast tumors, we examined the effects of RGS20 overexpression and knockdown on the cell mobility and adhesive properties of different human cancer cell lines, including cervical cancer HeLa, breast adenocarcinoma MDA-MB-231, and non-small cell lung carcinoma H1299 and A549 cells. Expression of RGS20 enhanced cell aggregation, migration, invasion and adhesion as determined by hanging drop aggregation, wound healing, transwell chamber migration and invasion assays. Conversely, shRNA-mediated knockdown of endogenous RGS20 impaired these responses. In addition, RGS20 elevated the expression of vimentin (a mesenchymal cell marker) but down-regulated the expression of E-cadherin, two indicators commonly associated with metastasis. These results suggest that the expression of RGS20 may promote metastasis of tumor cells.