The contribution of mRNA targeting to spatial protein localization in bacteria.

About 30% of all bacterial proteins execute their function outside of the cytosol and must be inserted into or translocated across the cytoplasmic membrane. This requires efficient targeting systems that recognize N-terminal signal sequences in client proteins and deliver them to protein transport complexes in the membrane. While the importance of these protein transport machineries for the spatial organization of the bacterial cell is well documented in multiple studies, the contribution of mRNA targeting and localized translation to protein transport is only beginning to emerge. mRNAs can exhibit diverse subcellular localizations in the bacterial cell and can accumulate at sites where new protein is required. This is frequently observed for mRNAs encoding membrane proteins, but the physiological importance of membrane enrichment of mRNAs and the consequences it has for the insertion of the encoded protein have not been explored in detail. Here, we briefly highlight some basic concepts of signal sequence-based protein targeting and describe in more detail strategies that enable the monitoring of mRNA localization in bacterial cells and potential mechanisms that route mRNAs to particular positions within the cell. Finally, we summarize some recent developments that demonstrate that mRNA targeting and localized translation can sustain membrane protein insertion under stress conditions when the protein-targeting machinery is compromised. Thus, mRNA targeting likely acts as a back-up strategy and complements the canonical signal sequence-based protein targeting.

mRNA targeting eliminates the need for the signal recognition particle during membrane protein insertion in bacteria.

Signal-sequence-dependent protein targeting is essential for the spatiotemporal organization of eukaryotic and prokaryotic cells and is facilitated by dedicated protein targeting factors such as the signal recognition particle (SRP). However, targeting signals are not exclusively contained within proteins but can also be present within mRNAs. By in vivo and in vitro assays, we show that mRNA targeting is controlled by the nucleotide content and by secondary structures within mRNAs. mRNA binding to bacterial membranes occurs independently of soluble targeting factors but is dependent on the SecYEG translocon and YidC. Importantly, membrane insertion of proteins translated from membrane-bound mRNAs occurs independently of the SRP pathway, while the latter is strictly required for proteins translated from cytosolic mRNAs. In summary, our data indicate that mRNA targeting acts in parallel to the canonical SRP-dependent protein targeting and serves as an alternative strategy for safeguarding membrane protein insertion when the SRP pathway is compromised.

Isoproterenol-Induced Cardiomyopathy Recovery Intervention: Amlexanox and Forskolin Enhances the Resolution of Catecholamine Stress-Induced Maladaptive Myocardial Remodeling.

The increasing incidence of stress-induced cardiomyopathy is due to the complexities of our modern-day lives, which constantly elicit stress responses. Herein, we aimed to explore the therapeutic potential of Amlexanox and Forskolin in promoting the recovery from stress-induced cardiomyopathy. Isoproterenol-induced cardiomyopathy (ICM) models were made, and the following treatment interventions were administered: 5% v/v DMSO as a placebo, Amlexanox (2.5 mg/100 g/day) treatment, Forskolin (0.5 mg/100 g/day), and Amlexanox and Forskolin combination, at their respective aforementioned dosages. The effects of Amlexanox and Forskolin treatment on ICM models were assessed by eletrocardiography and echocardiography. Also, using histological analysis and ELISA, their impact on myocardial architecture and inflammation were ascertained. ICM mice had excessive myocardial fibrosis, hypertrophy, and aggravated LVSDs which were accompanied by massive CD86+ inflammatory cells infiltration. Amlexanox treatment attenuated the myocardial hypertrophy, fibrosis, and inflammation and also slightly improved systolic functions. Meanwhile, forskolin treatment resulted in arrhythmias but significantly enhanced the resolution of myocardial fibrosis and inflammation. Intriguingly, Amlexanox and Forskolin combination demonstrated the most potency at promoting the recovery of the ICM from LVSD by attenuating maladaptive myocardial hypertrophy, fibrosis, and inflammatory responses. Our findings highlight the Amlexanox and Forskolin combination as a potential therapeutic intervention for enhancing cardiac function recovery from stress-induced cardiomyopathy by promoting the resolution of maladaptive cardiac remodeling.

Amlexanox and Forskolin Prevents Isoproterenol-Induced Cardiomyopathy by Subduing Cardiomyocyte Hypertrophy and Maladaptive Inflammatory Responses.

Chronic catecholamine stress (CCS) induces the occurrence of cardiomyopathy-pathological cardiac hypertrophy (PCH), which is characterized by left ventricular systolic dysfunction (LVSD). Recently, mounting evidence has implicated myocardial inflammation in the exacerbation of pathological cardiac remodeling. However, there are currently no well-defined treatment interventions or regimes targeted at both the attenuation of maladaptive myocardial hypertrophy and inflammation during CCS to prevent PCH. G protein-coupled receptor kinase 5 (GRK5) and adenylyl cyclases (ACs)-cAMP mediates both cardiac and inflammatory responses. Also, GRK5 and ACs are implicated in stress-induced LVSD. Herein, we aimed at preventing PCH during CCS via modulating adaptive cardiac and inflammatory responses by inhibiting GRK5 and/or stimulating ACs. Isoproterenol-induced cardiomyopathy (ICM) was modeled using 0.5 mg/100 g/day isoproterenol injections for 40 days. Alterations in cardiac and inflammatory responses were assessed from the myocardia. Similarities in the immunogenicity of cardiac troponin I (cTnI) and lipopolysaccharide under CCS were assessed, and Amlexanox (35 µM/ml) and/or Forskolin (10 µM/ml) were then employed in vitro to modulate adaptive inflammatory responses by inhibiting GRK5 or activating ACs-cAMP, respectively. Subsequently, Amlexanox (2.5 mg/100 g/day) and/or Forskolin (0.5 mg/100 g/day) were then translated into in vivo during CCS to modulate adaptive cardiac and inflammatory responses. The effects of Amlexanox and Forskolin on regulating myocardial systolic functions and inflammatory responses during CCS were ascertained afterward. PCH mice had excessive myocardial hypertrophy, fibrosis, and aggravated LVSD, which were accompanied by massive CD68+ inflammatory cell infiltrations. In vitro, Forskolin-AC/cAMP was effective than Amlexanox-GRK5 at downregulating proinflammatory responses during stress; nonetheless, Amlexanox and Forskolin combination demonstrated the most efficacy in modulating adaptive inflammatory responses. Individually, the translated Amlexanox and Forskolin treatment interventions were ineffective at subduing the pathological remodeling and sustaining cardiac function during CCS. However, their combination was potent at preventing LVSD during CCS by attenuating maladaptive myocardial hypertrophy, fibrosis, and inflammatory responses. The treatment intervention attained its potency mainly via Forskolin-ACs/cAMP-mediated modulation of cardiac and inflammatory responses, coupled with Amlexanox inhibition of GRK5 mediated maladaptive cascades. Taken together, our findings highlight the Amlexanox and Forskolin combination as a potential therapeutic intervention for preventing the occurrence of pathological cardiac hypertrophy during chronic stress.

Molecular mechanisms of circular RNAs, transforming growth factor-ß, and long noncoding RNAs in hepatocellular carcinoma.

At the heart of hepatocellular carcinoma (HCC) lies disruption of signaling pathways at the level of molecules, genes, and cells. Non-coding RNAs (ncRNAs) have been implicated in the disease progression of HCC. For instance, dysregulated expression of circular RNAs (circRNAs) has been observed in patients with HCC. As such, these RNAs are potential therapeutic targets and diagnostic markers for HCC. Long non-coding RNAs (lncRNAs), a type of ncRNA, have also been recognized to participate in the initiation and progression of HCC. Transforming growth factor-beta (TGF-ß) is another element which is now recognized to play crucial roles in HCC. It has been implicated in many biological processes such as survival, immune surveillance, and cell proliferation. In HCC, TGF-ß promotes disease progression by two mechanisms: an intrinsic signaling pathway and the extrinsic pathway. Through these pathways, it modulates various microenvironment factors such as inflammatory mediators and fibroblasts. An interesting yet-to-be resolved concept is whether the HCC-promoting role of TGF-ß pathways is limited to a subset of HCC patients or it is involved in the whole process of HCC development. This review summarizes recent advancements to highlight the roles of circRNAs, lncRNAs, and TGF-ß in HCC.

Pathological cardiac hypertrophy: the synergy of adenylyl cyclases inhibition in cardiac and immune cells during chronic catecholamine stress.

Response to stressors in our environment and daily lives is an adaptation conserved through evolution as it is beneficial in enhancing the survival and continuity of humans. Although stressors have evolved, the drastic physiological response they elicit still remains unchanged. The chronic secretion and circulation of catecholamines to produce physical responses when they are not required may result in pathological consequences which affect cardiac function drastically. This review seeks to point out the probable implication of chronic stress in inducing an inflammation disorder in the heart. We discussed the likely synergy of a G protein-independent stimuli signaling via ß2-adrenergic receptors in both cardiomyocytes and immune cells during chronic catecholamine stress. To explain this synergy, we hypothesized the possibility of adenylyl cyclases having a regulatory effect on G protein-coupled receptor kinases. This was based on the negative correlations they exhibit during normal cardiac function and heart failures. As such, the downregulation of adenylyl cyclases in cardiomyocytes and immune cells during chronic catecholamine stress enhances the expressions of G protein-coupled receptor kinases. In addition, we explain the maladaptive roles played by G protein-coupled receptor kinase and extracellular signal-regulated kinase in the synergistic cascade that pathologically remodels the heart. Finally, we highlighted the therapeutic potentials of an adenylyl cyclases stimulator to attenuate pathological cardiac hypertrophy (PCH) and improve cardiac function in patients developing cardiac disorders due to chronic catecholamine stress.

Estrogen regulation of cardiac cAMP-L-type Ca2+ channel pathway modulates sex differences in basal contraction and responses to ß2AR-mediated stress in left ventricular apical myocytes.

BACKGROUNDS/AIM: Male and female hearts have many structural and functional differences. Here, we investigated the role of estrogen (E2) in the mechanisms of sex differences in contraction through the cAMP-L-type Ca2+channel pathway in adult mice left ventricular (LV) apical myocytes at basal and stress state. METHODS: Isolated LV apical myocytes from male, female (Sham) and ovariectomised mice (OVX) were used to investigate contractility, Ca2+ transients and L-type Ca2+ channel (LTCC) function. The levels of ß2AR, intracellular cAMP, phosphodiesterase (PDE 3 and PDE 4), RyR2, PLB, SLN, and SERCA2a were compared among the experimental groups. RESULTS: We found that (1) intracellular cAMP, ICaL density, contraction and Ca2+ transient amplitudes were larger in Sham and OVX + E2 myocytes compared to male and OVX. (2) The mRNA expression of PDE 3 and 4 were lower in Sham and OVX + E2 groups compared with male and OVX groups. Treatment of myocytes with IBMX (100 µM) increased contraction and Ca2+ transient amplitude in both sexes and canceled differences between them. (3) ß2AR-mediated stress decreased cAMP concentration and peak contraction and Ca2+ transient amplitude only in male and OVX groups but not in Sham or OVX + E2 groups suggesting a cardioprotective role of E2 in female mice. (4) Pretreatment of OVX myocytes with GPR30 antagonist G15 (100 nM) abolished the effects of E2, but ERα and ERß antagonist ICI 182,780 (1 µM) did not. Moreover, activation of GPR30 with G1 (100 nM) replicated the effects of E2 on cAMP, contraction and Ca2+ transient amplitudes suggesting that the acute effects of E2 were mediated by GPR30 via non-genomic signaling. (5) mRNA expression of RyR2 was higher in myocytes from Sham than those of male while PLB and SLN were higher in male than Sham but no sex differences were observed in the mRNA of SERCA2a. CONCLUSION: Collectively, these results demonstrate that E2 modulates the expression of genes related to the cAMP-LTCC pathway and contributes to sex differences in cardiac contraction and responses to stress. We also show that estrogen confers cardioprotection against cardiac stress by non-genomic acute signaling via GPR30.

Decreased intracellular chloride promotes ADP induced platelet activation through inhibition of cAMP/PKA instead of activation of Lyn/PI3K/Akt pathway.

Decrease of chloride concentration contributes to cardiovascular diseases, however, whether decrease of chloride concentration is involved in platelet activation remains elusive. In the present study, we found that ACI patients had lower serum chloride which would be rescued after Aspirin administration. ADP induced chloride concentration reduction in platelets. Blockade of chloride channel prevented ADP-induced platelet adhesion, activation and aggregation, however, decreasing the extracellular chloride concentration promoted ADP-induced platelet adhesion and activation. Decrease of the extracellular chloride concentration facilitated the inactivation of Src family kinase Lyn, which was not involved in PI3K/Akt phosphorylation. Nevertheless, low chloride concentration promoted the production of platelet cytosol Gαi2 subunit. This subunit prevents AC from converting ATP into cAMP, which therefore, inhibited the phosphorylation of PKA to promote platelet activation. In conclusion, decreased intracellular chloride promotes ADP induced platelet activation through the Gαi2/cAMP/PKA pathway instead of the Lyn/PI3K/Akt signal pathway.

Low distribution of genes encoding virulence factors in Shigella flexneri serotypes 1b clinical isolates from eastern Chinese populations.

BACKGROUND: The ability of Shigella to invade, colonize, and eventually kill host cells is influenced by many virulence factors. However, there is no analysis of related genes in Jiangsu Province of China so far. Shigella flexneri was collected from 13 cities of Jiangsu Province through the provincial Centers for Disease Control (CDC) for analysis of distribution of major virulence genes (ipaH, ipaBCD, ial, virF, virB, sigA, set1A, sepA, sat, pic, set1B and sen) detected by PCR technology. RESULTS: A total of 545 isolates received were confirmed as S. flexneri which belongs to 11 serotypes of S. flexneri, among which serotype 2a was the most predominant (n = 223, 40.9%). All isolates were positive for ipaH gene, followed by sat (94.1%), sigA (78.9%), set1B (78.0%), pic (77.6%), set1A (74.5%), virF (64.8%), sepA (63.5%), sen (56.9%), ipaBCD (50.5%), ial (47.0%) and virB (47.0%). The presence of virulence genes in different serotypes was distinct. The existence of virulence genes of serotype 1b was generally lower than other serotype-the positive rate for virulence genes was between 0.0 and 14.1% except for ipaH and sat. In addition, virulence genes also fluctuated in different regions and at different times in Jiangsu province. The result of analysis on the relationship between virulence genes of S. flexneri showed that the existence of virulence genes of Shigella could be well represented by multiplex PCR combination ipaH + ial + set1A, which had a high clinical value. CONCLUSIONS: The present study was designed to explore the prevalence of 12 S. flexneri-associated virulence genes. The data showed high diversity of virulence genes with regard to periods, regions and serotypes in Jiangsu Province of China.