The ABL-MYC axis controls WIPI1-enhanced autophagy in lifespan extension.

Human WIPI ß-propellers function as PI3P effectors in autophagy, with WIPI4 and WIPI3 being able to link autophagy control by AMPK and TORC1 to the formation of autophagosomes. WIPI1, instead, assists WIPI2 in efficiently recruiting the ATG16L1 complex at the nascent autophagosome, which in turn promotes lipidation of LC3/GABARAP and autophagosome maturation. However, the specific role of WIPI1 and its regulation are unknown. Here, we discovered the ABL-ERK-MYC signalling axis controlling WIPI1. As a result of this signalling, MYC binds to the WIPI1 promoter and represses WIPI1 gene expression. When ABL-ERK-MYC signalling is counteracted, increased WIPI1 gene expression enhances the formation of autophagic membranes capable of migrating through tunnelling nanotubes to neighbouring cells with low autophagic activity. ABL-regulated WIPI1 function is relevant to lifespan control, as ABL deficiency in C. elegans increased gene expression of the WIPI1 orthologue ATG-18 and prolonged lifespan in a manner dependent on ATG-18. We propose that WIPI1 acts as an enhancer of autophagy that is physiologically relevant for regulating the level of autophagic activity over the lifespan.

Regulation of autophagosome biogenesis and mitochondrial bioenergetics by the cholesterol transport protein GRAMD1C.

Autophagosomes are crucial components of the cellular recycling machinery that form at endoplasmic reticulum (ER)-associated sites. As the autophagosome membrane is largely devoid of transmembrane proteins, autophagosome biogenesis is thought to be largely regulated by lipid transfer and lipid modifications, as well as membrane-associated proteins. While the membrane origin of autophagosomes and their lipid composition are still incompletely understood, previous studies have found the autophagosome membrane to be enriched in unsaturated fatty acids and have little cholesterol, suggesting that cholesterol removal is an integral step during autophagosome biogenesis. In our study, we demonstrate that short term cholesterol depletion leads to a rapid induction of autophagy and identify the ER-localized cholesterol transport protein GRAMD1C as a negative regulator of starvation-induced macroautophagy/autophagy. Abbreviations: ATG: autophagy related; ccRCC: clear cell renal cell carcinoma; ER: endoplasmic reticulum; GRAM: glucosyltransferases, RAB-like GTPase activators and myotubularins; GRAMD: GRAM domain containing; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MCBD: methyl-cyclodextrin; MTOR: mechanistic target of rapamycin kinase; VASt: VAD1 analog of StAR-related lipid transfer.

The cholesterol transport protein GRAMD1C regulates autophagy initiation and mitochondrial bioenergetics.

During autophagy, cytosolic cargo is sequestered into double-membrane vesicles called autophagosomes. The contributions of specific lipids, such as cholesterol, to the membranes that form the autophagosome, remain to be fully characterized. Here, we demonstrate that short term cholesterol depletion leads to a rapid induction of autophagy and a corresponding increase in autophagy initiation events. We further show that the ER-localized cholesterol transport protein GRAMD1C functions as a negative regulator of starvation-induced autophagy and that both its cholesterol transport VASt domain and membrane binding GRAM domain are required for GRAMD1C-mediated suppression of autophagy initiation. Similar to its yeast orthologue, GRAMD1C associates with mitochondria through its GRAM domain. Cells lacking GRAMD1C or its VASt domain show increased mitochondrial cholesterol levels and mitochondrial oxidative phosphorylation, suggesting that GRAMD1C may facilitate cholesterol transfer at ER-mitochondria contact sites. Finally, we demonstrate that expression of GRAMD family proteins is linked to clear cell renal carcinoma survival, highlighting the pathophysiological relevance of cholesterol transport proteins.

Increased cycling cell numbers and stem cell associated proteins as potential biomarkers for high grade human papillomavirus+ve pre-neoplastic cervical disease.

High risk (oncogenic) human papillomavirus (HPV) infection causes cervical cancer. Infections are common but most clear naturally. Persistent infection can progress to cancer. Pre-neoplastic disease (cervical intraepithelial neoplasia/CIN) is classified by histology (CIN1-3) according to severity. Cervical abnormalities are screened for by cytology and/or detection of high risk HPV but both methods are imperfect for prediction of which women need treatment. There is a need to understand the host virus interactions that lead to different disease outcomes and to develop biomarker tests for accurate triage of infected women. As cancer is increasingly presumed to develop from proliferative, tumour initiating, cancer stem cells (CSCs), and as other oncogenic viruses induce stem cell associated gene expression, we evaluated whether presence of mRNA (detected by qRT-PCR) or proteins (detected by flow cytometry and antibody based proteomic microarray) from stem cell associated genes and/or increased cell proliferation (detected by flow cytometry) could be detected in well-characterised, routinely collected cervical samples from high risk HPV+ve women. Both cytology and histology results were available for most samples with moderate to high grade abnormality. We found that stem cell associated proteins including human chorionic gonadotropin, the oncogene TP63 and the transcription factor SOX2 were upregulated in samples from women with CIN3 and that the stem cell related, cell surface, protein podocalyxin was detectable on cells in samples from a subset of women with CIN3. SOX2, TP63 and human gonadotrophin mRNAs were upregulated in high grade disease. Immunohistochemistry showed that SOX2 and TP63 proteins clearly delineated tumour cells in invasive squamous cervical cancer. Samples from women with CIN3 showed increased proliferating cells. We believe that these markers may be of use to develop triage tests for women with high grade cervical abnormality to distinguish those who may progress to cancer from those who may be treated more conservatively.

Corticotropin-releasing hormone receptors mediate opposing effects in cholestasis-induced liver cell apoptosis.

CRH receptors are expressed in human and rat liver. The current study investigated the biological role of the CRH system in the hepatocellular apoptotic process and aimed to reveal the responsible molecular mechanisms. Using a rat experimental model of common bile duct surgical ligation leading to obstructive jaundice and cholestasis, liver apoptosis was induced in the hepatic parenchyma as confirmed by the elevated expression of the early apoptotic neoepitope M30. This effect was reversed by administration of the nonselective CRH antagonist astressin but not by the selective CRH(2) antagonist astressin2B, suggesting that antagonism of the endogenous CRH(1) blocked the cholestasis-induced apoptotic mechanism. No effect was observed in the noncholestasis controls. In our experimental model, early and late apoptosis-preventing markers were induced in parallel to apoptosis; elevated gene transcript levels of the anti-apoptotic bcl-2 were found by real-time PCR in the first postoperative day and increased serum hepatocyte growth factor levels were measured by ELISA in the third postoperative day. Selective CRH(2) antagonism reversed the elevated expression of bcl-2 and hepatocyte growth factor, suggesting that this receptor type mediated antiapoptotic actions of the endogenous CRH system, opposing the preapoptotic ones mediated by CRH(1). In conclusion, the present study indicated that the CRH neuroendocrine system regulates cholestasis-induced apoptosis in the hepatic parenchyma via receptor-specific pathways. These data may contribute to better understanding of the CRH biology and its pathophysiological significance in the periphery.