GRP94 promotes muscle differentiation by inhibiting the PI3K/AKT/mTOR signaling pathway

The glucose-regulated endoplasmic reticulum chaperone protein 94 (GRP94) is required for many biological processes, such as secretion of immune factors and mesoderm induction. Here, we demonstrated that GRP94 promotes muscle differentiation in vitro and in vivo. Moreover, GRP94 inhibited the PI3K/AKT/mTOR signaling pathway. Using both in vitro and in vivo approaches, in myoblasts, we found that this inhibition resulted in reduced proliferation and increased differentiation. To further investigate the mechanism of GRP94-induced muscle differentiation, we used co-immunoprecipitation and proximity ligation assays and found that GRP94 interacted with PI3K-interacting protein 1 (Pik3ip1). The latter protein promoted muscle differentiation by inhibiting the PI3K/AKT/mTOR pathway. Furthermore, GRP94 was found to regulate Pik3ip1 expression. Finally, when Pik3ip1 expression was inhibited, GRP94-induced promotion of muscle differentiation was diminished. Taken together, our data demonstrated that GRP94 promoted muscle differentiation, mediated by Pik3ip1-dependent inhibition of the PI3K/AKT/mTOR signaling pathway.     

Altered signaling and cell cycle regulation in embryonal stem cells with a disruption of the gene for phosphoinositide 3-kinase regulatory subunit p85alpha

The p85alpha regulatory subunit of class I(A) phosphoinositide 3-kinases (PI3K) is derived from the Pik3r1 gene, which also yields alternatively spliced variants p50alpha and p55alpha. It has been proposed that excess monomeric p85 competes with functional PI3K p85-p110 heterodimers. We examined embryonic stem (ES) cells with heterozygous and homozygous disruptions in the Pik3r gene and found that wild type ES cells express virtually no monomeric p85alpha. Although, IGF-1-stimulated PI3K activity associated with insulin receptor substrates was unaltered in all cell lines, p85alpha-null ES cells showed diminished protein kinase B activation despite increased PI3K activity associated with the p85beta subunit. Furthermore, p85alpha-null cells demonstrated growth retardation, increased frequency of apoptosis, and altered cell cycle regulation with a G(0)/G(1) cell cycle arrest and up-regulation of p27(KIP), whereas signaling through CREB and MAPK was enhanced. These phenotypes were reversed by re-expression of p85alpha via adenoviral gene transfer. Surprisingly, all ES cell lines could be differentiated into adipocytes. In these differentiated ES cells, however, compensatory p85beta signaling was lost in p85alpha-null cells while increased signaling by CREB and MAPK was still observed. Thus, loss of p85alpha in ES cells induced alterations in IGF-1 signaling and regulation of apoptosis and cell cycle but no defects in differentiation. However, differentiated ES cells partially lost their ability for compensatory signaling at the level of PI3K, which may explain some of the defects observed in mice with homozygous deletion of the Pik3r1 gene.     

Positive and negative roles of p85 alpha and p85 beta regulatory subunits of phosphoinositide 3-kinase in insulin signaling

Class IA phosphoinositide (PI) 3-kinase is composed of a p110 catalytic subunit and a p85 regulatory subunit and plays a pivotal role in insulin signaling. To explore the physiological roles of two major regulatory isoforms, p85 alpha and p85 beta, we have established brown adipose cell lines with disruption of the Pik3r1 or Pik3r2 gene. Pik3r1-/- (p85 alpha-/-) cells show a 70% reduction of p85 protein and a parallel reduction of p110. These cells have a 50% decrease in PI 3-kinase activity and a 30% decrease in Akt activity, leading to decreased insulin-induced glucose uptake and anti-apoptosis. Pik3r2-/- (p85 beta-/-) cells show a 25% reduction of p85 protein but normal levels of p85-p110 and PI 3-kinase activity, supporting the fact that p85 is more abundant than p110 in wild type. p85 beta-/- cells, however, exhibit significantly increased insulin-induced Akt activation, leading to increased anti-apoptosis. Reconstitution experiments suggest that the discrepancy between PI 3-kinase activity and Akt activity is at least in part due to the p85-dependent negative regulation of downstream signaling of PI 3-kinase. Indeed, both p85 alpha-/- cells and p85 beta-/- cells exhibit significantly increased insulin-induced glycogen synthase activation. p85 alpha-/- cells show decreased insulin-stimulated Jun N-terminal kinase activity, which is restored by expression of p85 alpha, p85 beta, or a p85 mutant that does not bind to p110, indicating the existence of p85-dependent, but PI 3-kinase-independent, signaling pathway. Furthermore, a reduction of p85 beta specifically increases insulin receptor substrate-2 phosphorylation. Thus, p85 alpha and p85 beta modulate PI 3-kinase-dependent signaling by multiple mechanisms and transmit signals independent of PI 3-kinase activation.     

v-Crk activates the phosphoinositide 3-kinase/AKT pathway by utilizing focal adhesion kinase and H-Ras

v-Crk, an oncogene product of avian sarcoma virus CT10, efficiently transforms chicken embryo fibroblasts (CEF). We have recently reported that constitutive activation of the phosphoinositide 3-kinase (PI3K)/AKT pathway plays a critical role in the v-Crk-induced transformation of CEF. In the present study we investigated the molecular mechanism by which v-Crk activates the PI3K/AKT pathway. First, we found that v-Crk promotes the association of the p85 regulatory subunit of PI3K with focal adhesion kinase (FAK) by inducing the phosphorylation of the Y397 residue in FAK. This FAK phosphorylation needs activation of the Src family tyrosine kinase(s) for which the v-Crk SH2 domain is responsible. v-Crk was unable to activate the PI3K/AKT pathway in FAK-null cells, indicating the functional importance of FAK. In addition, we found that H-Ras is also required for the activation of the PI3K/AKT pathway. The v-Crk-induced activation of AKT was greatly enhanced by the overexpression of H-Ras or its guanine nucleotide exchange factor mSOS, which binds to the v-Crk SH3 domain, whereas a dominant-negative mutant of H-Ras almost completely suppressed this activation. Furthermore, we showed that v-Crk stimulates the interaction of H-Ras with the Ras binding domain in the PI3K p110 catalytic subunit. Our data indicated that the v-Crk-induced activation of PI3K/AKT pathway was cooperatively achieved by two distinct interactions. One is the interaction of p85 with tyrosine-phosphorylated FAK promoted by the v-Crk SH2 domain, and another is the interaction of p110 with H-Ras dictated by the v-Crk SH3 domain.     

Role of PI3K and AKT specific isoforms in ovarian cancer cell migration, invasion and proliferation through the p70S6K1 pathway

Ovarian cancer is the leading cause of death from gynecological malignancy for women. The amplification of the PI3K catalytic subunit (p110) and the lost function of PTEN are frequently detected in ovarian cancer cells. PI3K plays an important role in tumorigenesis. To specifically inhibit PI3K activity in ovarian cancer cells, we constructed small interfering RNA (siRNA) against p110. The expression of p110 siRNA significantly decreased cell migration, invasion, and proliferation compared to the siSCR control cells. The expression of p110 siRNA induced CDK inhibitor p27^KIP1 levels, and decreased levels of cyclin D1, CDK4, and phosphorylated retinoblastoma protein. PI3K transmits the mytogenic signal through AKT. AKT has three isoforms in the cells: AKT1, AKT2 and AKT3. We found that inhibition of AKT1 is sufficient to affect cell migration, invasion, and proliferation. Expression of AKT1 siRNA had a similar effect as p110 siRNA in the cells. We showed the roles of specific PI3K and AKT isoforms in the cells, which are important to understanding the mechanism of PI3K/AKT signaling in ovarian cancer cells. Both p110 and AKT1 siRNA-expressing cells decreased the activation of p70S6K1. Inhibition of p70S6K1 activity by its siRNA also decreased cell migration, invasion, and proliferation associated with the induction of p27^KIP1 levels, and with the inhibition of cell cycle-associated proteins including cyclin D1, CDK2, and phosphorylated retinoblastoma protein. This study demonstrates the important role of the PI3K/AKT/mTOR/p70S6K1 pathway in cell proliferation, migration, and invasion in ovarian cancer cells by using siRNA-mediated gene silencing as a reverse genetic method.     

Direct Association of Heat Shock Protein 20 (HSPB6) with Phosphoinositide 3-kinase (PI3K) in Human Hepatocellular Carcinoma: Regulation of the PI3K Activity

HSP20 (HSPB6), one of small heat shock proteins (HSPs), is constitutively expressed in various tissues and has several functions. We previously reported that the expression levels of HSP20 in human hepatocellular carcinoma (HCC) cells inversely correlated with the progression of HCC, and that HSP20 suppresses the growth of HCC cells via the AKT and mitogen-activated protein kinase signaling pathways. However, the exact mechanism underlying the effect of HSP20 on the regulation of these signaling pathways remains to be elucidated. To clarify the details of this effect in HCC, we explored the direct targets of HSP20 in HCC using human HCC-derived HuH7 cells with HSP20 overexpression. HSP20 proteins in the HuH7 cells were coimmunoprecipitated with the p85 regulatory subunit and p110 catalytic subunit of phosphoinositide 3-kinase (PI3K), an upstream kinase of AKT. Although HSP20 overexpression in HCC cells failed to affect the expression levels of PI3K, the activity of PI3K in the unstimulated cells and even in the transforming growth factor-α stimulated cells were downregulated by HSP20 overexpression. The association of HSP20 with PI3K was also observed in human HCC tissues in vivo. These findings strongly suggest that HSP20 directly associates with PI3K and suppresses its activity in HCC, resulting in the inhibition of the AKT pathway, and subsequently decreasing the growth of HCC.     

Class IA phosphoinositide 3-kinase regulates heart size and physiological cardiac hypertrophy

Class I(A) phosphoinositide 3-kinases (PI3Ks) are activated by growth factor receptors, and they regulate, among other processes, cell growth and organ size. Studies using transgenic mice overexpressing constitutively active and dominant negative forms of the p110alpha catalytic subunit of class I(A) PI3K have implicated the role of this enzyme in regulating heart size and physiological cardiac hypertrophy. To further understand the role of class I(A) PI3K in controlling heart growth and to circumvent potential complications from the overexpression of dominant negative and constitutively active proteins, we generated mice with muscle-specific deletion of the p85alpha regulatory subunit and germ line deletion of the p85beta regulatory subunit of class I(A) PI3K. Here we show that mice with cardiac deletion of both p85 subunits exhibit attenuated Akt signaling in the heart, reduced heart size, and altered cardiac gene expression. Furthermore, exercise-induced cardiac hypertrophy is also attenuated in the p85 knockout hearts. Despite such defects in postnatal developmental growth and physiological hypertrophy, the p85 knockout hearts exhibit normal contractility and myocardial histology. Our results therefore provide strong genetic evidence that class I(A) PI3Ks are critical regulators for the developmental growth and physiological hypertrophy of the heart.     

Ghrelin signaling in heart remodeling of adult obese mice

Ghrelin, an endogenous ligand of the growth hormone secretagogue receptor (GHS-R), has been suggested to be associated to obesity, insulin secretion, cardiovascular growth and homeostasis. GHS-R has been found in most of the tissues, and among the hormone action it is included the regulation of heart energy metabolism. Therefore, hypernutrition during early life leads to obesity, induces cardiac hypertrophy, compromises myocardial function, inducing heart failure in adulthood. We examined ghrelin signaling process in cardiac remodeling in these obese adult mice. The cardiomyocytes (cmy) of left ventricle were analyzed by light microscopy and stereology, content and phosphorilation of cardiac proteins: ghrelin receptor (growth hormone secretagogue receptor 1a, GHSR-1a), protein kinase B (AKT and pAKT), phosphatidil inositol 3 kinase (PI3K), AMP-activated protein kinase (AMPK and pAMPK) and actin were achieved by Western blotting. GHSR-1a gene expression was analyzed by Real Time-PCR. We observed hyperglycemia and higher liver and visceral fat weight in obese when compared to control group. Obese mice presented a marked increase in heart weight/tibia length, indicating an enlarged heart size or a remodeling process. Obese mice had increased GHSR-1a content and expression in the heart associated to PI3K content and increased AKT content and phosphorylation. In contrast, AMPK content and phosphorylation in heart was not different between experimental groups. Ghrelin plasma levels in obese group were decreased when compared to control group. Our data suggest that remodeled myocardial in adult obese mice overnourished in early life are associated with higher phosphorylation of GHSR-1a, PI3K and AKT but not with AMPK.     

Characterization of PI3K class IA isoforms with regulatory subunit p55alpha using a scintillation proximity assay

Differential activation of the phosphoinositide 3-kinase (PI3K)/AKT pathway has been linked to cancer. Activation occurs through gene amplification and activating mutations. High-frequency mutations in the gene encoding the p110α catalytic subunit of PI3K (PIK3CA) have been observed in a variety of tumors including colon, brain, breast, ovarian, and gastric. Inhibition of PI3K kinase activity may provide a specific way to treat multiple types of human cancer. A scintillation proximity assay (SPA) was developed to detect phosphatidylinositol 3-kinase catalytic activity. Using this assay format, steady-state kinetic parameters were compared for the PI3K class IA enzymes p110α, p110β, and p110δ, each coexpressed with the regulatory subunit p85α or splice variant p55α. Inhibition by the natural product wortmannin and LY294002 was detected with potencies consistent with alternate assay formats. Other biochemical assay formats have been described for phosphoinositide 3-kinases but each has its unique limitations. The simple, inexpensive, sensitive high-throughput nature of the SPA format has advanced our knowledge of isoform-specific enzymology and will facilitate the discovery of novel PI3K inhibitors.     

G1 cell cycle progression and the expression of G1 cyclins are regulated by PI3K/AKT/mTOR/p70S6K1 signaling in human ovarian cancer cells

Ovarian cancer is one of the most common cancers among women. Recent studies demonstrated that the gene encoding the p110alpha catalytic subunit of phosphatidylinositol 3-kinase (PI3K) is frequently amplified in ovarian cancer cells. PI3K is involved in multiple cellular functions, including proliferation, differentiation, antiapoptosis, tumorigenesis, and angiogenesis. In this study, we demonstrate that the inhibition of PI3K activity by LY-294002 inhibited ovarian cancer cell proliferation and induced G(1) cell cycle arrest. This effect was accompanied by the decreased expression of G(1)-associated proteins, including cyclin D1, cyclin-dependent kinase (CDK) 4, CDC25A, and retinoblastoma phosphorylation at Ser(780), Ser(795), and Ser(807/811). Expression of CDK6 and beta-actin was not affected by LY-294002. Expression of the cyclin kinase inhibitor p16(INK4a) was induced by the PI3K inhibitor, whereas steady-state levels of p21(CIP1/WAF1) were decreased in the same experiment. The inhibition of PI3K activity also inhibited the phosphorylation of AKT and p70S6K1, but not extracellular regulated kinase 1/2. The G(1) cell cycle arrest induced by LY-294002 was restored by the expression of active forms of AKT and p70S6K1 in the cells. Our study shows that PI3K transmits a mitogenic signal through AKT and mammalian target of rapamycin (mTOR) to p70S6K1. The mTOR inhibitor rapamycin had similar inhibitory effects on G(1) cell cycle progression and on the expression of cyclin D1, CDK4, CDC25A, and retinoblastoma phosphorylation. These results indicate that PI3K mediates G(1) progression and cyclin expression through activation of an AKT/mTOR/p70S6K1 signaling pathway in the ovarian cancer cells.     

Stem cell factor/c-kit up-regulates cyclin D3 and promotes cell cycle progression via the phosphoinositide 3-kinase/p70 S6 kinase pathway in spermatogonia

Stem cell factor (SCF)/c-kit plays an important role in the regulation of hematopoiesis, melanogenesis, and spermatogenesis. In the testis, the SCF/c-kit system is believed to regulate germ cell proliferation, meiosis, and apoptosis. Studies with type A spermatogonia in vivo and in vitro have indicated that SCF induces DNA synthesis and proliferation. However, the signaling pathway for this function of SCF/c-kit has not been elucidated. We now demonstrate that SCF activates phosphoinositide 3-kinase (PI3-K) and p70 S6 kinase (p70S6K) and that rapamycin, a FRAP/mammalian target of rapamycin-dependent inhibitor of p70S6K, completely inhibited bromodeoxyuridine incorporation induced by SCF in primary cultures of spermatogonia. SCF induced cyclin D3 expression and phosphorylation of the retinoblastoma protein through a pathway that is sensitive to both wortmannin and rapamycin. Furthermore, AKT, but not protein kinase C-zeta, is used by SCF/c-kit/PI3-K to activate p70S6K. Dominant negative AKT-K179M completely abolished p70S6K phosphorylation induced by the constitutively active PI3-K catalytic subunit p110. Constitutively active v-AKT highly phosphorylated p70S6K, which was totally inhibited by rapamycin. Thus, SCF/c-kit uses a rapamycin-sensitive PI3-K/AKT/p70S6K/cyclin D3 pathway to promote spermatogonial cell proliferation.     

Molecular Balance between the Regulatory and Catalytic Subunits of Phosphoinositide 3-Kinase Regulates Cell Signaling and Survival

Class Ia phosphoinositide (PI) 3-kinase is a central component in growth factor signaling and is comprised of a p110 catalytic subunit and a regulatory subunit, the most common family of which is derived from the p85alpha gene (Pik3r1). Optimal signaling through the PI 3-kinase pathway depends on a critical molecular balance between the regulatory and catalytic subunits. In wild-type cells, the p85 subunit is more abundant than p110, leading to competition between the p85 monomer and the p85-p110 dimer and ineffective signaling. Heterozygous disruption of Pik3r1 results in increased Akt activity and decreased apoptosis by insulin-like growth factor 1 (IGF-1) through up-regulated phosphatidylinositol (3,4,5)-triphosphate production. Complete depletion of p85alpha, on the other hand, results in significantly increased apoptosis due to reduced PI 3-kinase-dependent signaling. Thus, a reduction in p85alpha represents a novel therapeutic target for enhancing IGF-1/insulin signaling, prolongation of cell survival, and protection against apoptosis.     

14-3-3 proteins regulate glycogen synthase 3beta phosphorylation and inhibit cardiomyocyte hypertrophy

14-3-3 proteins are dimeric phophoserine-binding molecules that participate in important cellular processes such as cell proliferation, cell-cycle control and the stress response. In this work, we report that several isoforms of 14-3-3s are expressed in neonatal rat cardiomyocytes. To understand their function, we utilized a general 14-3-3 peptide inhibitor, R18, to disrupt 14-3-3 functions in cardiomyocytes. Cardiomyocytes infected with adenovirus-expressing YFP-R18 (AdR18) exhibited markedly increased protein synthesis and atrial natriuretic peptide production and potentiated the responses to norepinephrine stimulation. This response was blocked by the pretreatment with LY294002, a phosphoinositide 3-kinase (PI3K) inhibitor. Consistent with a role of PI3K in the R18 effect, R18 induced phosphorylation of a protein cloned from the vakt oncogene of retrovirus AKT8 (Akt - also called protein kinase B, PKB) at Ser473 and glycogen synthase 3beta (GSK3beta) at Ser9, but not extracellular signal-regulated kinase 1/2 (ERK1/2). AdR18-induced PKB and GSK3beta phosphorylation was completely blocked by LY294002. In addition, a member of the nuclear factor of activated T cells (NFAT) family, NFAT3, was converted into faster mobility forms and translocated into the nucleus upon the treatment of AdR18. These results suggest that 14-3-3s inhibits cardiomyocytes hypertrophy through regulation of the PI3K/PKB/GSK3beta and NFAT pathway.     

Activation of peroxisome proliferator-activated receptor-alpha prevents glycogen synthase 3beta phosphorylation and inhibits cardiac hypertrophy

Activation of peroxisome proliferator-activated receptor-alpha (PPAR-alpha) has been recently reported to inhibit vascular inflammatory response and prevent cardiac hypertrophy. However, it is unclear how the activation of PPAR-alpha regulates hypertrophic response. In the present study, we found that application of fenofibrate and overexpression of PPAR-alpha inhibited endothelin-1 (ET-1)-induced phosphorylation of protein kinase B (Akt) at Ser473 and glycogen synthase kinase3beta (GSK3beta) at Ser9, and prevented ET-1-induced nuclear translocation of NFATc4 in cardiomyocytes. Moreover, co-immunoprecipitation studies showed that fenofibrate strongly induced the association of nuclear factor of activated T cells (NFATc4) with PPAR-alpha. These results suggest that activation of PPAR-alpha inhibits ET-1-induced cardiac hypertrophy through regulating PI3K/Akt/GSK3beta and NFAT signaling pathways.     

Identification and characterization of the autophosphorylation sites of phosphoinositide 3-kinase isoforms beta and gamma

Class I phosphoinositide 3-kinases (PI3Ks) are bifunctional enzymes possessing lipid kinase activity and the capacity to phosphorylate their catalytic and/or regulatory subunits. In this study, in vitro autophosphorylation of the G protein-sensitive p85-coupled class I(A) PI3K beta and p101-coupled class I(B) PI3K gamma was examined. Autophosphorylation sites of both PI3K isoforms were mapped to C-terminal serine residues of the catalytic p110 subunit (i.e. serine 1070 of p110 beta and serine 1101 of p110 gamma). Like other class I(A) PI3K isoforms, autophosphorylation of p110 beta resulted in down-regulated PI3K beta lipid kinase activity. However, no inhibitory effect of p110 gamma autophosphorylation on PI3K gamma lipid kinase activity was observed. Moreover, PI3K beta and PI3K gamma differed in the regulation of their autophosphorylation. Whereas p110 beta autophosphorylation was stimulated neither by G beta gamma complexes nor by a phosphotyrosyl peptide derived from the platelet-derived growth factor receptor, autophosphorylation of p110 gamma was significantly enhanced by G beta gamma in a time- and concentration-dependent manner. In summary, we show that autophosphorylation of both PI3K beta and PI3K gamma occurs in a C-terminal region of the catalytic p110 subunit but differs in its regulation and possible functional consequences, suggesting distinct roles of autophosphorylation of PI3K beta and PI3K gamma.     

Regulation of L-type inward calcium channel activity by captopril and angiotensin II via the phosphatidyl inositol 3-kinase pathway in cardiomyocytes from volume-overload hypertrophied rat hearts

Heart failure can be caused by pro-hypertrophic humoral factors such as angiotensin II (Ang II), which regulates protein kinase activities. The intermingled responses of these kinases lead to the early compensated cardiac hypertrophy, but later to the uncompensated phase of heart failure. We have shown that although beneficial, cardiac hypertrophy is associated with modifications in ion channels that are mainly mediated through mitogen-activated protein (MAP) kinase and phosphatidylinositol 3-kinase (PI3K) activation. This study evaluates the control of L-type Ca(2+) current (I(Ca,L)) by the Ang II/PI3K pathway in hypertrophied ventricular myocytes from volume-overload rats using the perforated patch-clamp technique. To assess activation of the I(Ca,L) in cardiomyocytes, voltages of 350 ms in 10 mV increments from a holding potential of -85 mV were applied to cardiocytes, with a pre-pulse to -45 mV for 300 ms. Volume overload-induced hypertrophy reduces I(Ca,L), whereas addition of Ang II alleviates the hypertrophic-induced decrease in a PI3K-dependent manner. Acute administration of Ang II (10(-6) mol/L) to normal adult cardiomyocytes had no effect; however, captopril reduced their basal I(Ca,L). In parallel, captopril regressed the hypertrophy and inverted the Ang II effect on I(Ca,L) seemingly through a PI3K upstream effector. Thus, it seems that regression of cardiac hypertrophy by captopril improved I(Ca,L) partly through PI3K.     

The role of phosphatidylinositol 3'-kinase and its downstream signals in erbB-2-mediated transformation

We previously demonstrated that erbB-2-overexpressing human mammary epithelial (HME) cells exhibit several transformed phenotypes including growth factor independence, anchorage-independent growth, motility, and invasiveness. Because phosphatidylinositol 3'-kinase (PI3K) is a major target of erbB-2 activation, we tested the contribution that PI3K and its downstream signaling pathways make to these phenotypes. Utilizing a constitutively active form of PI3K, p110CAAX, we show that PI3K can mediate most phenotypes observed in erbB-2-overexpressing cells. To identify pathways leading from PI3K to specific phenotypes, we expressed constitutively active AKT or PTEN in erbB-2-overexpressing cells or in HME cells. HME cells expressing constitutively active AKT were growth factor independent, anchorage independent and motile, but not invasive. PTEN expression blocked erbB-2-mediated invasion but none of the other phenotypes. Rottlerin blocked invasion induced by p110CAAX and erbB-2, suggesting that protein kinase C delta (PKC-delta) is the downstream effector of PI3K responsible for the invasive capacity of the cells. Consistent with these observations, phospho-AKT remained detectable in erbB-2 cells treated with LY294002 or expressing exogenous PTEN, but was abolished by treatment with the p38MAP kinase inhibitor SB202190. Thus, both PI3K-dependent and p38MAP kinase-dependent pathways lead to activation of AKT, and activation of PKC-delta, via PI3K, mediates invasion.     

Class III PI3K‐mediated prolonged activation of autophagy plays a critical role in the transition of cardiac hypertrophy to heart failure

Pathological cardiac hypertrophy often leads to heart failure. Activation of autophagy has been shown in pathological hypertrophic hearts. Autophagy is regulated positively by Class III phosphoinositide 3‐kinase (PI3K). However, it is unknown whether Class III PI3K plays a role in the transition of cardiac hypertrophy to heart failure. To address this question, we employed a previously established cardiac hypertrophy model in heat shock protein 27 transgenic mice which shares common features with several types of human cardiomyopathy. Age‐matched wild‐type mice served as control. Firstly, a prolonged activation of autophagy, as reflected by autophagosome accumulation, increased LC3 conversion and decreased p62 protein levels, was detected in hypertrophic hearts from adaptive stage to maladaptive stage. Moreover, morphological abnormalities in myofilaments and mitochondria were presented in the areas accumulated with autophagosomes. Secondly, activation of Class III PI3K Vacuolar protein sorting 34 (Vps34), as demonstrated by upregulation of Vps34 expression, increased interaction of Vps34 with Beclin‐1, and deceased Bcl‐2 expression, was demonstrated in hypertrophic hearts from adaptive stage to maladaptive stage. Finally, administration with Wortmaninn, a widely used autophagy inhibitor by suppressing Class III PI3K activity, significantly decreased autophagy activity, improved morphologies of intracellular apartments, and most importantly, prevented progressive cardiac dysfunction in hypertrophic hearts. Collectively, we demonstrated that Class III PI3K plays a central role in the transition of cardiac hypertrophy to heart failure via a prolonged activation of autophagy in current study. Class III PI3K may serve as a potential target for the treatment and management of maladaptive cardiac hypertrophy.