Biology subjects

Cohen, J. C.

Publications and source records attributed to Cohen, J. C..

4 recordsLinked to original sources

bioRxiv2026

TM6SF2 binds cholesterol, interacts with apolipoprotein B, and promotes hepatic lipid secretion

A missense variant in TM6SF2 (transmembrane 6 superfamily member 2, TM6SF2E167K) is a major risk factor for steatotic liver disease1, while protecting against coronary artery disease2. TM6SF2 is a polytopic resident protein of the smooth endoplasmic reticulum (ER) and ER-Golgi intermediate compartment that promotes lipidation of hepatic ApoB-containing lipoproteins before secretion into the circulation. Here, we used cryo-electron microscopy (cryo-EM) to determine the structures of TM6SF2 and TM6SF2E167K at 3.64 [A] and 3.58 [A] resolution, respectively. TM6SF2 comprises 10 transmembrane helices that bind a single cholesterol molecule within a transmembrane cavity. The protein assembles into homodimers and homotetramers that interact with ApoB. Structural and biochemical analyses show that the E167K substitution reduces cholesterol binding and ApoB interaction without disrupting overall protein structure. Expression of wild-type, but not mutant, TM6SF2 restores hepatic triglyceride secretion in TM6SF2-deficient hepatocytes. Together, these findings establish the first structural framework for the bulk lipidation step in hepatic lipoprotein biogenesis, the principal pathway for hepatic triglyceride and cholesterol export into the circulation.

cell biology↗

bioRxiv2026

Dual Control of LDL-cholesterol Levels by ANGPTL3 and ANGPTL8

BACKGROUNDInactivation of ANGPTL3 (angiopoietin-like protein 3, A3) is a proven therapeutic strategy for lowering plasma lipid levels independently of the LDL receptor (LDLR), yet the optimal approach to inactivate A3 remains unclear. A3 is proteolytically cleaved and circulates as full-length (A3-FL), N-terminal (A3-Nter) and C-terminal (A3-Cter) fragments. The specific contribution of each form of A3, and of its paralog, ANGPTL8 (A8), in modulating circulating levels of ApoB-Containing Lipoproteins (ABCLs) remain poorly defined. Clarifying these relationships will inform next-generation A3-directed therapies. METHODSWe performed liver perfusion studies to directly compare the number and composition of VLDL particles secreted from mice with and without A3. To amplify effects on cholesterol metabolism, we generated Ldlr-/- mice expressing wildtype A3 (A3-WT), A3-FL or A3-Nter, with or without co-expression of A8, and analyzed plasma lipids, circulating A3 and A8 complexes, and intravascular lipase activities. Complementary in vitro assays and structural modeling were used to assess relative endothelial lipase (EL) inhibition by A3 alone or in complex with A8. RESULTSLiver perfusion studies revealed that A3 inactivation does not alter the rates of hepatic secretion of VLDL in wildtype or Ldlr-/- mice. Inactivation of A8 alone lowered plasma LDL-cholesterol (C) levels by [~]20%, an effect dependent upon the expression of both EL and A3. Maximal inhibition of lipoprotein lipase (LPL) required co-expression of A8 plus both A3-FL and A3-Nter, indicating that A3 cleavage, in addition to A8 expression, is essential for maximal LPL inhibition. In contrast, A8 expression, but not A3 cleavage, was required for optimal EL inhibition. CONCLUSIONSA8 acts in concert with A3 to differentially modulate LPL- and EL-mediated lipolysis, which antagonizes hepatic clearance of newly-secreted atherogenic ABCLs. This mechanistic framework refines our understanding of A3-targeted lipid lowering and highlights the therapeutic potential of dual A3- plus A8-directed strategies to treat dyslipidemia and prevent atherosclerotic cardiovascular disease. Clinical perspectiveO_ST_ABSWhat is new?C_ST_ABSO_LIInactivation of A3 lowers circulating ABCL levels without altering hepatic secretion rates of VLDL-ApoB or -TG. C_LIO_LIProteolytic cleavage of A3 is required for maximal inhibition of LPL. C_LIO_LIInactivation of A8 lowers LDL-C levels through an A3- and EL-dependent, but LDLR-independent, mechanism. C_LI What are the clinical implications?O_LICombining A8 inhibition with A3-inactivating therapies offers a strategy to achieve greater reduction in LDL-C levels and atherosclerotic cardiovascular risk. C_LI

molecular biology↗

bioRxiv2025

Tissue-specific regulation of PNPLA3 promotes lipid remodeling in response to dietary and temperature stress

Background & AimsPNPLA3(I148M) is the strongest genetic risk factor for steatotic liver disease (SLD), but its functional role and tissue-specific regulation remain unclear. In mice, PNPLA3 is abundant in liver, yet undetectable in adipose depots. Here, we characterize the molecular mechanisms underlying these tissue-specific differences in PNPLA3 expression in mice to clarify its functional role and link to SLD risk. MethodsPnpla3 mRNA and PNPLA3 protein levels were quantified in liver and adipose depots of fasted and refed mice at 30{degrees}C and 6{degrees}C. Signaling pathways regulating PNPLA3 expression in adipocytes were examined using adrenergic agonists and pathway-specific modulators. Translation and proteasomal inhibitors were used during adrenergic stimulation to investigate the discordance between Pnpla3 mRNA and protein levels. Relationship between PNPLA3 levels and triglyceride (TG) fatty acid composition was also assessed. ResultsAt thermoneutrality, feeding strongly increased PNPLA3 levels in liver but it remained undetectable in adipose tissue of mice. Conversely, cold exposure or {beta}3-adrenergic stimulation had no effect on hepatic PNPLA3, but increased PNPLA3 >19-fold in brown adipose tissue (BAT), despite causing a >75% reduction in Pnpla3 mRNA, indicating robust post-translational regulation. In BAT, adrenergic signaling via cAMP/PKA and PI3K/AKT elevated PNPLA3 by reducing proteasomal degradation. PNPLA3 expression correlated with depletion of TG-long-chain polyunsaturated fatty acids (TG-LCPUFAs) in both liver and BAT, consistent with a role in lipid remodeling. ConclusionsThese findings reveal striking tissue- and context-specific regulation of PNPLA3, but a conserved association between its expression and TG-LCPUFAs levels, suggesting that PNPLA3 modulates lipid remodeling in response to metabolic stress and that disrupting this function may contribute to SLD susceptibility. Impact and implicationsDespite being the strongest genetic risk factor for SLD, PNPLA3s physiological role remains unclear. Using mouse models, this study reveals that PNPLA3 is regulated in a tissue-specific manner in response to feeding and cold exposure, thereby promoting remodeling of cellular lipids to adapt to dietary and environmental challenges. The localization of PNPLA3 action and its tissue-specific regulation are directly relevant to hepatologists and metabolic researchers aiming to understand its influence on intracellular lipid composition and its effects on disease susceptibility. Moreover, modulation of PNPLA3 turnover--and its impact on LCPUFAs remodeling--emerges as a potential therapeutic strategy for regulating lipid homeostasis in SLD. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=191 SRC="FIGDIR/small/684800v2_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@ee6ac7org.highwire.dtl.DTLVardef@a45d74org.highwire.dtl.DTLVardef@f39929org.highwire.dtl.DTLVardef@cc826b_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIPNPLA3 is regulated in a highly tissue-specific manner in mice. C_LIO_LIIn liver, feeding-but not cold exposure-induces PNPLA3 primarily through transcriptional mechanisms. C_LIO_LIIn adipose tissue, cold exposure-but not feeding-induces PNPLA3 through post-transcriptional mechanisms. C_LIO_LIIn adipose tissue, cold exposure increases PNPLA3 despite a reduction in Pnpla3 mRNA. C_LIO_LIPNPLA3 remodels lipids in liver and adipose tissue to maintain lipid homeostasis, a process disrupted in SLD. C_LI

physiology↗

bioRxiv2024

PNPLA3(148M) Promotes Hepatic Steatosis by Interfering with Triglyceride Hydrolysis Through a Gain-of-Function Mechanism

Background & AimsPNPLA3(148M) (patatin-like phospholipase domain-containing protein 3) is the most impactful genetic risk factor for steatotic liver disease (SLD), thus motivating a search for therapeutic modulators of its expression. A key unresolved issue is whether PNPLA3(148M) confers a loss- or gain-of-function. Here we used multiple approaches to further test the hypothesis that PNPLA3 causes steatosis by sequestering ABHD5 (/{beta} hydrolase domain containing protein 5), the cofactor of ATGL (adipose TG lipase), thus limiting mobilization of hepatic triglyceride (TG). MethodsWe quantified the physical interactions between ABHD5 and PNPLA3/ATGL in cultured hepatocytes using NanoBiT complementation assays. Immunocytochemistry was used to compare the relative binding of PNPLA3 and ATGL to ABHD5 and to determine if PNPLA3 must associate with lipid droplets (LDs) to inhibit ATGL. Adenoviruses and adeno-associated viruses were used to express PNPLA3 in liver-specific Atgl-/- mice and ABHD5 in livers of Pnpla3148M/M mice, respectively. We used purified recombinant proteins to compare the TG hydrolytic activities of PNPLA3 and ATGL in the presence and absence of ABHD5. ResultsABHD5 interacted preferentially with PNPLA3 relative to ATGL in cultured hepatocytes and in vitro, with no differences observed between PNPLA3(WT) or PNPLA3(148M). PNPLA3(148M)-associated inhibition of TG hydrolysis required localization of PNPLA3 to LDs and the presence of ATGL. Finally, overexpression of ABHD5 reversed the hepatic steatosis in Pnpla3M/M mice. ConclusionsThese findings support the premise that PNPLA3(148M) promotes hepatic steatosis by accumulating on LDs and inhibiting ATGL-mediated lipolysis in an ABHD5-dependent manner. Our results predict that reducing, rather that increasing PNPLA3 expression will be the best strategy to treat PNPLA3(148M)-associated SLD. Impact and implicationsSteatotic liver disease (SLD) is a common complex disorder associated with both environmental and genetic risk factors. PNPLA3(148M) is the most impactful genetic risk factor for SLD and yet its pathogenic mechanism remains controversial. Here we provide evidence that PNPLA3(148M) promotes triglyceride (TG) accumulation by sequestering ABHD5, thus limiting its availability to activate ATGL. Although the substitution of methionine for isoleucine reduces the TG hydrolytic activity of PNPLA3, the loss-of-function is only indirectly related to the steatotic effect of the variant. Here we provide evidence that PNPLA3(148M) confers a gain-of-function by interfering with ATGL-mediated TG hydrolysis. These findings have implications for the design of potential PNPLA3-based therapies. Reducing, rather than increasing, PNPLA3 levels is predicted to reverse steatosis in susceptible individuals. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=106 SRC="FIGDIR/small/606015v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@1dd7acaorg.highwire.dtl.DTLVardef@1a869dborg.highwire.dtl.DTLVardef@1bde743org.highwire.dtl.DTLVardef@99effc_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIABHD5 binds preferentially to PNPLA3 relative to ATGL. C_LIO_LIPNPLA3(WT) and PNPLA3(148M) compete similarly for binding and inhibition of ATGL. C_LIO_LIABHD5 activates the triglyceride lipase activity of PNPLA3, as well as ATGL. C_LIO_LIThe steatotic effect of PNPLA3(148M) requires expression of ATGL. C_LIO_LIOverexpression of ABHD5 can rescue the steatosis associated with PNPLA3(148M). C_LI

cell biology↗