2A Protein Research Articles

Overcoming methicillin resistance by methicillin-resistant Staphylococcus aureus: Computational evaluation of napthyridine and oxadiazoles compounds for potential dual inhibition of PBP-2a and FemA proteins

Abstract The treatment of the various infections caused by Staphylococcus aureus has become challenging due to the evolving resistance against current therapeutics. In this study, the potentials of napthyridine and oxadiazole derivatives to serve as dual inhibitors of penicillin-binding protein 2a (PBP-2a) and FemA protein, which are crucial to resistance to methicillin-based drugs by S. aureus, were evaluated using molecular modeling techniques. Seventy-two compounds were subjected to molecular docking against the proteins, and the hit compounds were subjected to drug-likeness evaluation and in silico pharmacokinetics prediction. The compounds with good safety profiles were subjected to a 250-ns molecular dynamics (MD) simulation and other relevant analyses based on the MD trajectories. Five hit compounds were selected based on their high affinity for the targets as evidenced by their docking scores ranging from −8.6 to −10.1 kcal/mol for PBP-2a and −9.6 to −9.9 kcal/mol for FemA. These compounds also passed Lipinski’s rule of five evaluation with no violation and possessed high human intestinal absorption potential, showcasing their potential as orally administered therapeutic agents. However, three of the compounds were potential mutagens. MD simulation revealed that the final two compounds maintained stable interactions with the target proteins over 250 ns, with minimal deviations and fluctuations. Hydrogen bond stability and energy decomposition analysis further confirmed the strong binding affinity of the hit compounds compared to the control drug, methicillin. Conclusively, the compounds with the CID “135964525” and “44130718” are worthy of further experimental validation in the development of potential inhibitors of PBP-2a and FemA.

Protein phosphatase 6 promotes stemness of colorectal cancer cells.

Colorectal cancer (CRC) remains a significant global health concern, demanding a more profound comprehension of its molecular foundations for the development of improved therapeutic strategies. This study aimed to elucidate the role of protein phosphatase 6 (PP6), a member of the type 2A protein phosphatase family, in CRC. Protein phosphatase 6 functions as a heterotrimer with a catalytic subunit (PP6c), regulatory subunits (PP6Rs; PP6R1, PP6R2, and PP6R3), and scaffold subunits (ANKRD28, ANKRD44, and ANKRD52). Elevated PP6c expression has been identified in CRC tissues compared to normal mucosa, aligning with its potential involvement in CRC pathogenesis. PP6c knockdown resulted in decreased colony-forming ability and invivo proliferation of various CRC cell lines. Transcriptome analysis revealed that PP6c knockdown resulted in altered expression of genes associated with cancer stemness. Notably, the PP6c-PP6R3 complex is a key player in regulating cancer stem cell (CSC) markers. Additionally, increased PP6c expression was observed in CSC-like cells induced by sphere formation, implicating the role of PP6c in CSC maintenance. This study highlights the role of PP6c in CRC and suggests that it is a potential therapeutic target disrupting a pathway critical for CRC progression and stem cell maintenance.

Effects of Perioperative Zinc Supplementation on Copper Circulating Levels and Expression of Metallothionein and Copper Antioxidant Chaperone-1 in Leukocytes in Patients Undergoing CABG Surgery.

The use of zinc supplement may have a negative effect on copper status. The objective of this study was to evaluate the effect of zinc and vitamin E supplementation on copper and zinc biomarkers in patients undergoing coronary artery bypass graft (CABG) surgery. The study was an add-on project to a previously published randomized controlled trial (NCT05402826) on patients undergoing CABG surgery. Patients in the zinc-vitamin E group (n = 40) received oral zinc (120 mg) and vitamin E (1200 international units) 1 day before surgery, followed by 30 mg of zinc and 200 units of vitamin E per day until 21 days after surgery, while those in the control group (n = 38) received placebo. Plasma levels of copper, ceruloplasmin, superoxide dismutase (SOD) activity, as well as leukocyte gene expression of metallothionein 2A (MT2A) and antioxidant protein 1 (ATOX1), were determined 3 and 21 days after surgery. The plasma copper level in the zinc-vitamin E group was significantly lower than the placebo group on the 3rd postoperative day, but no significant between-group differences were observed on day 21. Plasma ceruloplasmin concentration and SOD activity were not different. Relative mRNA expression of leukocyte MT2A was increased at both times (days 3 and 21 in the zinc-vitamin E group compared to placebo, but ATOX1 expression was not affected. Although the plasma copper level was transiently decreased early after surgery in the zinc-vitamin E group, considering the lack of change in other copper biomarkers, it seems that the use of zinc supplements at the dose used in the present study does not have a significant negative effect on the role of intracellular copper.

Selection of texture-associated biomarkers in chilled and iced grass carp (Ctenopharyngodon idella) fillets via DIA-based proteomics

Consumers care about the texture of fresh fish flesh, but a rapid quantitative analytical method for this has not been properly established. In this study, texture-associated biomarkers were selected by DIA-based proteomics for possible future application. Results indicated a significant decline in texture and moisture characteristics with extended storage under chilled and iced conditions, and flesh quality was categorized into three intervals. A total of 8 texture-associated biomarkers were identified in the chilled storage group, and 3 distinct ones in the iced storage group. Biomarkers were further refined based on their expression levels. Isobutyryl-CoA dehydrogenase, mitochondrial and [Phosphatase 2A protein]-leucine-carboxy methyltransferase were identified as effective texture-associated biomarkers for chilled fish, and Staphylococcal nuclease domain-containing protein 1 for iced fish. This study provided suitable proteins as indicators of fresh fish flesh texture, which could help establish a rapid and convenient texture testing method in future studies.

Glycyrrhizic Acid Nanoparticles Subside the Activity of Methicillin-Resistant Staphylococcus aureus by Suppressing PBP2a.

Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA) are classified as high-risk infections that can lead to death, particularly among older individuals. Nowadays, plant nanoparticles such as glycyrrhizic acid are recognized as efficient bactericides against a wide range of bacterial strains. Recently, scientists have shown interest in plant extract nanoparticles, derived from natural sources, which can be synthesized into nanomaterials. Interestingly, glycyrrhizic acid is rich in antioxidants as well as antibacterial agents, and it exhibits no adverse effects on normal cells. In this study, glycyrrhizic acid nanoparticles (GA-NPs) were synthesized using the hydrothermal method and characterized through physicochemical techniques such as UV-visible spectrometry, DLS, zeta potential, and TEM. The antimicrobial activity of GA-NPs was investigated through various methods, including MIC assays, anti-biofilm activity assays, ATPase activity assays, and kill-time assays. The expression levels of mecA, mecR1, blaR1, and blaZ genes were measured by quantitative RT-qPCR. Additionally, the presence of the penicillin-binding protein 2a (PBP2a) protein of S. aureus and MRSA was evaluated by a Western blot assay. The results emphasized the fabrication of GA nanoparticles in spherical shapes with a diameter in the range of 40-50 nm. The data show that GA nanoparticles exhibit great bactericidal effectiveness against S. aureus and MRSA. The treatment with GA-NPs remarkably reduces the expression levels of the mecA, mecR1, blaR1, and blaZ genes. PBP2a expression in MRSA was significantly reduced after treatment with GA-NPs. Overall, this study demonstrates that glycyrrhizic acid nanoparticles have potent antibacterial activity, particularly against MRSA. This research elucidates the inhibition mechanism of glycyrrhizic acid, which involves the suppressing of PBP2a expression. This work emphasizes the importance of utilizing plant nanoparticles as effective antimicrobial agents against a broad spectrum of bacteria.

HSPA8 Chaperone Complex Drives Chaperone-Mediated Autophagy Regulation in Acute Promyelocytic Leukemia Cell Differentiation

Introduction: Acute myeloid leukemia (AML) is a cancer of the hematopoietic system characterized by hyperproliferation of undifferentiated cells of the myeloid lineage. While most of AML therapies are focused toward tumor debulking, all-trans retinoic acid (ATRA) induces neutrophil differentiation in the AML subtype acute promyelocytic leukemia (APL). Macroautophagy has been extensively investigated in the context of various cancers and is often dysregulated in AML where it can have context-dependent pro- or anti-leukemogenic effects. On the contrary, the implications of chaperone-mediated autophagy (CMA) on the pathophysiology of diseases are still being explored and its role in AML remains elusive. Methods: We took advantage of human AML primary samples and databases to analyze CMA gene expression and activity. Furthermore, we used ATRA-sensitive (NB4) and -resistant (NB4-R1) APL cells to further dissect a potential function for CMA in ATRA-mediated neutrophil differentiation. NB4-R1 cells are unique in that they do respond to retinoic acid transcriptionally but do not mature in response to retinoid signaling alone unless maturation is triggered by adding cyclic adenosine monophosphate. Results: Here, we report that CMA-related mRNA transcripts are significantly higher expressed in immature hematopoietic cells as compared to neutrophils, contrasting the macroautophagy gene expression patterns. Accordingly, lysosomal degradation of an mCherry-KFERQ CMA reporter decreases during ATRA-induced differentiation of APL cells. On the other hand, using NB4-R1 cells we found that macroautophagy flux primed ATRA-resistant NB4-R1 cells to differentiate upon ATRA treatment but reduced the association of lysosome-associated membrane protein type 2A (LAMP-2A) and heat shock protein family A (Hsp70) member 8 (HSPA8), necessary for complete neutrophil maturation. Accordingly, depletion of HSPA8 attenuated CMA activity and facilitated APL cell differentiation. In contrast, maintaining high CMA activity by ectopic expression of LAMP-2A impeded APL differentiation. Conclusion: Overall, our findings suggest that APL neutrophil differentiation requires CMA inactivation and that this pathway predominantly depends on HSPA8 and is possibly assisted by other co-chaperones.

The N-Terminal Region of Cucumber Mosaic Virus 2a Protein Is Involved in the Systemic Infection in Brassica juncea.

Brassica juncea belongs to the Brassicaceae family and is used as both an oilseed and vegetable crop. As only a few studies have reported on the cucumber mosaic virus (CMV) in B. juncea, we conducted this study to provide a basic understanding of the B. juncea and CMV interactions. B. juncea-infecting CMV (CMV-Co6) and non-infecting CMV (CMV-Rs1) were used. To identify the determinants of systemic infection in B. juncea, we first constructed infectious clones of CMV-Co6 and CMV-Rs1 and used them as pseudo-recombinants. RNA2 of CMV was identified as an important determinant in B. juncea because B. juncea were systemically infected with RNA2-containing pseudo-recombinants; CMV-Co6, R/6/R, and R/6/6 were systemically infected B. juncea. Subsequently, the amino acids of the 2a and 2b proteins were compared, and a chimeric clone was constructed. The chimeric virus R/6Rns/R6cp, containing the C-terminal region of the 2a protein of CMV-Rs1, still infects B. juncea. It is the 2a protein that determines the systemic CMV infection in B. juncea, suggesting that conserved 160G and 214A may play a role in systemic CMV infection in B. juncea.

The catalytic subunit of type 2A protein phosphatase negatively regulates conidiation and melanin biosynthesis in Setosphaeria turcica

Northern corn leaf blight caused by Setosphaeria turcica is a major fungal disease responsible for significant reductions in maize yield worldwide. Eukaryotic type 2A protein phosphatase (PP2A) influences growth and virulence in a number of pathogenic fungi, but little is known about its roles in S. turcica. Here, we functionally characterized S. turcica StPP2A-C, which encodes the catalytic C subunit of StPP2A. StPP2A-C deletion slowed colony growth, conidial germination, and appressorium formation but increased conidiation, melanin biosynthesis, glycerol content, and disease lesion size on maize. These effects were associated with expression changes in genes related to calcium signaling, conidiation, laccase activity, and melanin and glycerol biosynthesis, as well as changes in intra- and extracellular laccase activity. A pull-down screen for candidate StPP2A-c interactors revealed an interaction between StPP2A-c and StLac1. Theoretical modeling and yeast two-hybrid experiments confirmed that StPP2A-c interacted specifically with the copper ion binding domain of StLac1 and that Cys267 of StPP2A-c was required for this interaction. StPP2A-C expression thus appears to promote hyphal growth and reduce pathogenicity in S. turcica, at least in part by altering melanin synthesis and laccase activity; these insights may ultimately support the development of novel strategies for biological management of S. turcica.

Short Communication: Mosquito Histone 2A Protein Facilitate Japanese Encephalitis Virus Infection in the Mosquito.

Japanese encephalitis virus is mainly prevalent in the tropical and subtropical regions of Asia and Oceania. Through immunoprecipitation-mass spectrometry analysis using monoclonal antibodies targeting JEV E protein, we found that mosquito Histone 2A protein could bind to JEV particles. The binding of H2A and JEV was detected in the salivary gland and supernatant of mosquito cells. Furthermore, RNA interference experiments in vitro and in vivo confirmed that H2A protein promotes JEV infection in mosquitoes. In summary, we found that mosquito H2A is a factor that supports JEV infection and can potentially facilitate cross-species transmission of JEV.

Tracing synaptic loss in Alzheimer's brain with SV2A PET-tracer UCB-J.

Synaptic loss is an early prominent feature of Alzheimer's disease (AD). The recently developed novel synaptic vesicle 2A protein (SV2A) PET-tracer UCB-J has shown great promise in tracking synaptic loss in AD. However, there have been discrepancies between the findings and a lack of mechanistic insight. Here we report the first extensive pre-clinical validation studies for UCB-J in control (CN; n=11) and AD (n=11) brains using a multidimensional approach of post-mortem brain imaging techniques, radioligand binding, and biochemical studies. We demonstrate that UCB-J could target SV2A protein with high specificity and depict synaptic loss at synaptosome levels in AD brain regions compared to CNs. UCB-J showed highest synaptic loss in AD hippocampus followed in descending order by frontal cortex, temporal cortex, parietal cortex, and cerebellum. 3H-UCB-J large brain-section autoradiography and cellular/subcellular fractions binding studies indicated potential off-target interaction with phosphorylated tau (p-tau) species in AD brains, which could have subsequent clinical implications for imaging studies. Synaptic positron emission tomography (PET)-tracer UCB-J could target synaptic vesicle 2A protein (SV2A) with high specificity in Alzheimer's disease (AD) and control brains. Synaptic PET-tracer UCB-J could depict synaptic loss at synaptosome levels in AD brain regions compared to control. Potential off-target interaction of UCB-J with phosphorylated tau (p-tau) species at cellular/subcellular levels could have subsequent clinical implications for imaging studies, warranting further investigations.

Development of Stable Infectious cDNA Clones of Tomato Black Ring Virus Tagged with Green Fluorescent Protein.

Tomato black ring virus (TBRV) is a member of the Nepovirus genus in the Secoviridae family, which infects a wide range of important crop species worldwide. In this work, we constructed four cDNA infectious clones of the TBRV tagged with the green fluorescent protein (TBRV-GFP), which varied in (i) the length of the sequences flanking the GFP insert, (ii) the position of the GFP insert within the RNA2 polyprotein, and (iii) the addition of a self-cutting 2A protein. The presence of the GFP coding sequence in infected plants was verified by RT-PCR, while the infectivity and stability of the constructs were verified by mechanical inoculation of the host plants. The systemic spread of TBRV-GFP within plants was observed under UV light at a macroscopic level, monitoring GFP-derived fluorescence in leaves, and at a microscopic level using confocal microscopy. The obtained clones are a valuable tool for future studies of TBRV-host interactions, virus biology, and the long-term monitoring of its distribution in infected plants.

The role of the encephalomyocarditis virus type 1 proteins L and 2A in the inhibition of the synthesis of cellular proteins and the accumulation of viral proteins during infection

Infection of cells with encephalomyocarditis virus type 1 (EMCV-1, Cardiovirus A: Picornaviridae) is accompanied by suppression of cellular protein synthesis. The main role in the inhibition of cellular translation is assigned to the L and 2A «security» proteins. The mechanism of the possible influence of the L protein on cellular translation is unknown. There are hypotheses about the mechanism of influence of 2A protein on the efficiency of cap-dependent translation, which are based on interaction with translation factors and ribosome subunits. However, the available experimental data are contradictory, obtained using different approaches, and do not form a unified model of the interaction between the L and 2A proteins and the cellular translation machinery. To study the role of L and 2A «security» proteins in the suppression of translation of cellular proteins and the efficiency of translation and processing of viral proteins in infected cells. Mutant variants of EMCV-1 were obtained to study the properties of L and 2A viral proteins: Zfmut, which has a defective L; Δ2A encoding a partially deleted 2A; Zfmut&Δ2A containing mutations in both proteins. Translational processes in infected cells were studied by Western-blot and the pulse method of incorporating radioactively labeled amino acids (14C) into newly synthesized proteins, followed by radioautography. The functional inactivation of the 2A protein does not affect the inhibition of cellular protein synthesis. A direct correlation was found between the presence of active L protein and specific inactivation of cellular protein synthesis at an early stage of viral infection. Nonspecific suppression of the translational processes of the infected cell, accompanied by phosphorylation of eIF2α, occurs at the late stage of infection. Partial removal of the 2A protein from the EMCV-1 genome does not affect the development of this process, while inactivation of the L protein accelerates the onset of complete inhibition of protein synthesis. Partial deletion of the 2A disrupts the processing of viral capsid proteins. Suppression of L protein functions leads to a decrease in the efficiency of viral translation. A study of the role of EMCV-1 L and 2A proteins during the translational processes of an infected cell, first performed using infectious viral pathogens lacking active L and 2A proteins in one experiment, showed that 2A protein is not implicated in the inhibition of cellular translation in HeLa cells; L protein seems to play an important role not only in the specific inhibition of cellular translation but also in maintaining the efficient synthesis of viral proteins; 2A protein is involved not only in primary but also in secondary processing of EMCV-1 capsid proteins.

Catharanthus roseus (L.) G. Don counteracts the ampicillin resistance in multiple antibiotic-resistant Staphylococcus aureus by downregulation of PBP2a synthesis.

It is essential to revisit the global biodiversity, search for ethnopharmacologically relevant plants, and unveil their untapped potential to overcome the complications associated while treating infections triggered by multiple antibiotic-resistant Staphylococcus aureus. Catharanthus roseus (L.) G. Don of the Apocynaceae family is a medicinal plant used for remedial purposes against infectious diseases from ancient times. In this study, we intended to evaluate the mechanism by which the ethanolic extract of C. roseus root (EECRR) causes the reversal of ampicillin resistance in S. aureus. To achieve this goal, we have stained EECRR-treated S. aureus with acridine orange, analysed DNA damage by comet assay, and studied the alteration of plasmid band pattern and expression of penicillin-binding protein 2a (PBP2a) protein. Experiments revealed better S. aureus killing efficiency of EECRR at its minimum inhibitory concentration (MIC) doses due to DNA damage and reducing plasmid band intensities along with a decline in the expression of PBP2a in EECRR-treated cells at half-MIC dose. EECRR proved to be an efficient growth inhibitor of S. aureus that reduces the expression of PBP2a. Therefore, EECRR can also render ampicillin-resistant S. aureus susceptible to the antibiotic.

5-Methoxytryptophan alleviates atrial structural remodeling in ibrutinib-associated atrial fibrillation

BackgroundIbrutinib is an effective and well-tolerated treatment for B-cell lymphomas but is associated with an increased risk of atrial fibrillation (AF) by altering the structure of the atrium. 5-Methoxytryptophan (5-MTP) inhibits inflammatory and fibrotic processes. This study aimed to determine the effects and mechanisms of 5-MTP on the underlying mechanisms of AF caused by ibrutinib. MethodsThe effect of 5-MTP on ibrutinib-related AF was investigated in male Sprague Dawley rats using echocardiographic, electrophysiological, immunofluorescent, Masson staining, and molecular analyses. RusultsThe ibrutinib+5-MTP group showed (1) a lower incidence and shorter duration of AF and accelerated atrial conduction; (2) a decreased left atrial mass and left atrial diameter; (3) decreased myocardial fibrosis in the left atrium; (4) lower atrial inflammation; (5) increased sarcoplasmic reticulum Ca2+-ATPase 2a protein expression, decreased phosphorylation of phospholamban at Thr17, and decreased sodium/calcium exchanger 1 protein expression and phosphorylation of ryanodine receptor 2 at S2814; and (6) decreased phosphorylation of CaMKII expression. 5-MTP treatment markedly activated the PI3K-Akt signaling. Inhibiting PI3K-Akt signaling significantly reversed the protective effect of 5-MTP on ibrutinib-related AF. ConclusionsThese findings suggest that 5-MTP administration decreases the vulnerability of ibrutinib-related AF mainly caused by ameliorated maladaptive left atrial remodeling and dysregulation of calcium handling proteins. Mechanistically, 5-MTP treatment markedly enhanced the activation of cardiac PI3K-Akt signaling.

Repurposing Amphotericin B: anti-microbial, molecular docking and molecular dynamics simulation studies suggest inhibition potential of Amphotericin B against MRSA

Amphotericin B (AMPH) is an anti-fungal drug and this study, for the first time as best of our knowledge, reports the repurposing of the Amphotericin B. The drug was found to show significant antibacterial potential revealed by antimicrobial screening, molecular docking, and mode of action analysis targeting Penicillin Binding Protein 2a (PBP 2a protein) which is target of β-lactam drugs and is involved in cell wall synthesis. Mode of action analysis showed the drug to have hydrophobic and hydrophilic interactions with both C-terminal, trans-peptidase and non-penicillin binding domain of the protein. Additionally, to evaluate the impact of ligand binding on the protein's conformational dynamics, molecular dynamics (MD) simulations were used. Comparative Dynamical flexibility (RMSF) and Dynamics Cross Correlation (DCCM) followed by MD simulations revealed the complex formation significantly effecting structural dynamics of the enzyme significantly in the non-penicillin binding domain (327–668) and slightly in trans peptidase domain. Radius of gyration assessment further showed ligand binding also decreasing over all compactness of protein. Secondary structure analysis indicated the complex formation changing the conformational integrity in non-penicillin binding domain. Hydrogen bond analysis and MMPBSA, free energy of calculations followed by MD simulations, also complemented the antimicrobial and molecular docking revelations suggesting Amphotericin B to have substantial antibacterial potential.

Ca2+ handling remodelling in a porcine model of right ventricular dysfunction

Increased afterload is the primary pathophysiologic mechanism for right ventricular failure (RVF), which is the most critical determinant of survival in patients with pulmonary hypertension (PH). The cellular and molecular processes involved in the RV remodeling, including Ca2+ handling as an instrumental actor in the heart, remain incompletely understood. We gain insights into the cellular Ca2+ signaling in a large animal model of RV failure, the chronic thromboembolic pulmonary hypertension (CTEPH) piglet model, to develop specific therapy targeting RVF. CTEPH piglet model consisted of a primary left pulmonary artery ligation followed by weekly embolization in the right lower lobe leading to PH and progressive RV hypertrophy and failure. After six weeks, the piglet model replicated all the features of human CTEPH: increased pulmonary vascular resistance, mean pulmonary arterial pressure (PAP), and RV wall thickness associated with RV dysfunction as assessed by the reduced tricuspid annular plane systolic excursion (TAPSE). RV histological analyses revealed hypertrophied cardiomyocytes and aberrant fibrosis. At the cell level, RV myocytes from CTEPH piglets presented reduced L-type Ca2+ currents and CACNA1C mRNA, lower and slower [Ca2+]i transients, decreased sarcoplasmic reticulum Ca2+ content, and cell shortening to myocytes from sham piglets. This was related to reduce Sarco/Endoplasmic Reticulum Ca2+-ATPase isoform 2a (SERCA2a) protein expression. We found that store-operated Ca2+ entry is increased in hypertrophied RV myocytes, which is correlated with a de novo expression of the SR Ca2+ sensor STIM1L (long stromal interacting molecule 1). Finally, we found that Ca2+ sparks are larger and longer in RV myocytes from CTEPH piglets, whereas their frequencies and amplitude were unchanged compared to myocytes from sham. This was associated with profound disorganization of T tubules and ryanodine receptors (RyR) clustering without changes in RyR phosphorylation status. Our data highlight a cellular Ca2+ cycling remodeling that participates to the pathogenesis of RVF in CTEPH piglets.

Cadmium exposure induces a sex-dependent decline in left ventricular cardiac function

AimsCadmium exposure is a worldwide problem that has been linked to the development of cardiovascular disease. This study aimed to elucidate mechanistic details of chronic cadmium exposure on the structure and function of the heart. Main methodsMale and female mice were exposed to cadmium chloride (CdCl2) via drinking water for eight weeks. Serial echocardiography and blood pressure measurements were performed. Markers of hypertrophy and fibrosis were assessed, along with molecular targets of Ca2+-handling. Key findingsMales exhibited a significant reduction in left ventricular ejection fraction and fractional shortening with CdCl2 exposure, along with increased ventricular volume at end-systole, and decreased interventricular septal thickness at end-systole. Interestingly, no changes were detected in females. Experiments in isolated cardiomyocytes revealed that CdCl2-induced contractile dysfunction was also present at the cellular level, showing decreased Ca2+ transient and sarcomere shortening amplitude with CdCl2 exposure. Further mechanistic investigation uncovered a decrease in sarco/endoplasmic reticulum Ca2+-ATPase 2a (SERCA2a) protein expression and phosphorylated phospholamban levels in male hearts with CdCl2 exposure. SignificanceThe findings of our novel study provide important insight into how cadmium exposure may act as a sex-specific driver of cardiovascular disease, and further underscore the importance of reducing human exposure to cadmium.

Cross-Ethnic Variant Screening and Burden Analysis of PTPA in Parkinson's Disease.

Recently, homozygous variants in PTPA were identified as the disease cause for two pedigrees with early-onset parkinsonism and intellectual disability. Although the initial link between PTPA and parkinsonism has been established, further replication was still necessary. To evaluate the genetic role of PTPA in Parkinson's disease (PD). We analyzed rare variants of PTPA in cohorts of Asian and European ancestries (Ncase = 2743, Ncontrol = 8177) with whole-exome sequencing, and further explored the functional effect of the target variant. One patient with early-onset PD from a consanguineous family carried the homozygous variant p.Met329Val, while her parents and elder sister with heterozygous p.Met329Val were healthy. This patient developed minor cognitive decline within 1 year, with a Montreal Cognitive Assessment (MoCA) score dropping from 28 to 25. Functional exploration with overexpression studies suggested that this variant was associated with decreased protein phosphatase 2A (PTPA) protein level by affecting protein stability, but not mRNA expression. These results have broadened the mutation spectrum of PTPA, and paved the way for further research into the role of PTPA in PD. © 2023 International Parkinson and Movement Disorder Society.

HIGD2A silencing impairs hepatocellular carcinoma growth via inhibiting mitochondrial function and the MAPK/ERK pathway

BackgroundThe Hypoxia inducible gene domain family member 2A (HIGD2A) protein is indispensable for the assembly of the mitochondrial respiratory supercomplex, which has been implicated in cell proliferation and cell survival under hypoxic conditions. Because the liver has a naturally low oxygen microenvironment, the role of HIGD2A in the development of hepatocellular carcinoma (HCC) remains largely unknown.MethodsGene expression data and clinical information were obtained from multiple public databases. A lentivirus-mediated gene knockdown approach was conducted to explore the function and mechanism of HIGD2A activity in HCC cells. In vivo and in vitro assays were performed to investigate the biological roles of HIGD2A.ResultsHIGD2A was overexpressed in HCC tissues and cell lines and was associated with a worse prognosis. Silencing HIGD2A expression significantly attenuated cell proliferation and migration, caused S-phase cell cycle arrest, and decreased tumor formation in nude mice. Mechanistically, HIGD2A depletion greatly decreased cellular ATP levels by disrupting mitochondrial ATP production. Moreover, HIGD2A knockdown cells displayed impaired mitochondrial function, such as mitochondrial fusion, increased expression of the mitochondrial stress response protein, and decreased oxygen consumption. Furthermore, knockdown of HIGD2A markedly attenuated the activation of the MAPK/ERK pathway.ConclusionsHIGD2A promoted liver cancer cell growth by fueling mitochondrial ATP synthesis and activating the MAPK/ERK pathway, suggested that targeting HIGD2A may represent a new strategy for HCC therapy.

Aging decreases docosahexaenoic acid transport across the blood-brain barrier in C57BL/6J mice.

Nutrients are actively taken up by the brain via various transporters at the blood-brain barrier (BBB). A lack of specific nutrients in the aged brain, including decreased levels of docosahexaenoic acid (DHA), is associated with memory and cognitive dysfunction. To compensate for decreased brain DHA, orally supplied DHA must be transported from the circulating blood to the brain across the BBB through transport carriers, including major facilitator superfamily domain-containing protein 2a (MFSD2A) and fatty acid-binding protein 5 (FABP5) that transport esterified and non-esterified DHA, respectively. Although it is known that the integrity of the BBB is altered during aging, the impact of aging on DHA transport across the BBB has not been fully elucidated. We used 2-, 8-, 12-, and 24-month-old male C57BL/6 mice to evaluate brain uptake of [14C]DHA, as the non-esterified form, using an in situ transcardiac brain perfusion technique. Primary culture of rat brain endothelial cells (RBECs) was used to evaluate the effect of siRNA-mediated MFSD2A knockdown on cellular uptake of [14C]DHA. We observed that the 12- and 24-month-old mice exhibited significant reductions in brain uptake of [14C]DHA and decreased MFSD2A protein expression in the brain microvasculature compared with that of the 2-month-old mice; nevertheless, FABP5 protein expression was up-regulated with age. Brain uptake of [14C]DHA was inhibited by excess unlabeled DHA in 2-month-old mice. Transfection of MFSD2A siRNA into RBECs decreased the MFSD2A protein expression levels by 30% and reduced cellular uptake of [14C]DHA by 20%. These results suggest that MFSD2A is involved in non-esterified DHA transport at the BBB. Therefore, the decreased DHA transport across the BBB that occurs with aging could be due to age-related down-regulation of MFSD2A rather than FABP5.