Biology subjects

THIAM, A. R.

Publications and source records attributed to THIAM, A. R..

8 recordsLinked to original sources

bioRxiv2026

ER cholesteryl ester phase separation underlies switch-like cholesterol sensing

Cellular lipid homeostasis requires mechanisms that detect subtle changes in lipid abundance and trigger rapid, coordinated responses. The INSIG/SCAP/SREBP2 pathway provides a central feedback system linking endoplasmic reticulum (ER) cholesterol levels to the transcriptional control of cholesterol genes, yet the origin of its remarkable cooperative switch mechanism remains unclear. Here we identify cholesterol esterification as a physical mechanism that amplifies sterol sensing and generates switch-like pathway regulation. We show that cholesteryl oleate (CE), produced by SOAT1 and opposed by NCEH1-mediated hydrolysis, undergoes a cooperative phase transition within the ER membrane to form transient CE-rich domains. These lipid assemblies create a threshold-dependent platform that concentrates SCAP and promotes formation of the SCAP-INSIG retention complex, thereby coupling ER lipid organization to SREBP inhibition. Because CE domain formation is nucleation-driven, variations in cholesterol availability are converted into an abrupt transition between distinct membrane states. Perturbing CE metabolism uncouples cholesterol abundance from pathway activity: SOAT1 inhibition prevents CE domain formation, releases SCAP from the ER, and activates SREBP2 despite cholesterol sufficiency, whereas NCEH1 inhibition stabilizes the domains and reduces SREBP2 activation under cholesterol-limiting conditions. Thus, the balance between esterification and hydrolysis determines a membrane physical state that serves as the functional output sensed by the cholesterol regulatory machinery. Our findings reveal ER lipid phase transitions as a general principle for creating ultrasensitive control in cellular homeostasis and establish cholesterol esterification as an active regulatory process rather than a passive storage pathway.

cell biology↗

bioRxiv2026

Triglyceride/Cholesterol Ester Ratio Encodes Lipid Droplet Size and Diversity

Lipid droplet (LD) heterogeneity is a hallmark of their pathophysiological relevance. This is especially evident when cells sequester toxic cholesterol by converting it into cholesterol esters (CEs) within LDs. Since CEs can form liquid crystals (LCs), it remains unclear how such ordered structures are accommodated within the inherently dynamic environment of LDs. Here, we show that the fluidizing properties of triglycerides (TGs) help CE incorporation and influence LD growth and heterogeneity. Seipin, the key regulator of TG-LD formation and size, does not significantly impact CE-rich LD size. Instead, the CE/TG ratio, the sequence of neutral lipid deposition, and the activity of diacylglycerol acyltransferases, especially DGAT2, determine whether LDs enlarge, remain fluid, or lock into LC phases. We found that the LC phase resists LD ripening and acts as a kinetic barrier to lipid entry. Lastly, we observe that perilipins, the most abundant LD surface proteins, differentially target CE-rich LDs: Plin3 and 5 are excluded from them, whereas Plin2 and 4 are favored. These findings highlight the CE/TG ratio as a key organizing principle of lipid storage and LD function, with immediate relevance for diseases linked to sterol accumulation.

cell biology↗

bioRxiv2024

Steric Repulsion Counteracts ER-to-Lipid Droplet Protein Movement

Lipid droplets (LDs) are uniquely shaped organelles consisting of a neutral lipid core surrounded by a phospholipid monolayer, continuous with the cytosolic leaflet of the endoplasmic reticulum (ER). The dynamics and function of LDs are closely tied to their proteome composition, which is subject to dynamic remodeling. Key proteins essential for LD biology relocate from the ER to LDs, yet the mechanisms governing their movement and accumulation in LDs remain poorly understood. Here, we developed an innovative ex vivo tool to quantify and classify ER proteins based on their affinity for LDs. We found a broad spectrum of ER-to-LD partitioning affinities. We identified steric hindrance as a key factor in regulating ER-to-LD protein transfer, where proteins with only slightly higher LD affinity can effectively displace those with lower affinity from the LD surface. Consistent with this model, we observed that differentiation of 3T3 pre-adipocytes into adipocytes involves extensive remodeling of ER proteins targeting LDs, with Plin1--a high-affinity LD protein--becoming predominantly recruited and excluding other ER proteins. These findings highlight lateral protein-protein exclusion as a fundamental mechanism in shaping the LD proteome, providing new insights into LD biogenesis and function.

biophysics↗

bioRxiv2024

Seipin Regulates Caveolin-1 Trafficking and Organelle Crosstalk

Caveolin-1 (CAV1), the main structural component of caveolae, is essential in various biological processes, including mechanotransduction, lipid metabolism, and endocytosis1-4. Deregulation of CAV1 dynamics is linked to various pathologies, including cellular senescence, cancer, insulin resistance, and lipodystrophy5-9. However, mechanisms regulating CAV1 trafficking and function remain poorly understood. Here, we show that seipin, a crucial lipid droplet (LD) biogenesis factor10, modulates CAV1 trafficking. Deletion of seipin resulted in the accumulation of saturated lipids, leading to ceramide and sphingomyelin overproduction, which disrupted the membrane order of the trans-Golgi network (TGN). In seipin deficiency, CAV1 location to the plasma membrane (PM) was impaired, reducing caveolae. Instead, CAV1 accumulated in TGN and late endosome compartments, which fused with LDs and delivered the protein. In wild-type (WT) cells, this process was minimal but significantly enhanced by treatment with palmitate, ceramide, or Stearoyl-CoA desaturase-1 (SCD1) inhibition. Conversely, in seipin-deficient cells, inhibiting Fatty Acid Synthase (FASN) or overexpressing SCD1 restored CAV1 localization to the PM and reduced its accumulation in LDs. Our findings reveal that seipin controls the funneling of palmitate toward glycerolipids synthesis and storage in LDs versus conversion to ceramides in the ER. This balance is crucial to cellular protein trafficking by controlling the TGN membrane order. Therefore, our study identifies seipin as a critical regulator of cellular lipid metabolism, protein trafficking, and organelle homeostasis. These findings shed light on the processes regulating CAV1 trafficking and show that convergent pathophysiological mechanisms associated with defects in CAV1 and seipin contribute to metabolic disorders, including insulin resistance and lipodystrophies11-14.

cell biology↗

bioRxiv2024

α-Synuclein acts as a cholesteryl-ester sensor on lipid droplets regulating organelle size and abundance.

While aggregated alpha-Synuclein (Syn) is commonly associated with Parkinsons disease, its physiological function as a membrane-binding protein is poorly understood. Here, we show that endogenous Syn binds lipid droplets (LDs) in multiple human cell lines and in stem cell-derived dopaminergic neurons. LD-binding encompasses Syn residues 1-100, which masks their detection by immunofluorescence microscopy, probably explaining the scarcity of similar observations in earlier studies. Syn-LD interactions are highly temperature-sensitive and selective for cholesteryl-ester-rich LDs. They promote the formation of Syn multimers that dissociate from LDs at non-permissive temperatures. Syn remains LD-bound throughout starvation-induced lipolysis, whereas siRNA-knockdown diminishes LD abundance and compromises cell viability upon nutrient depletion, without affecting LD biosynthesis. Reciprocally, excess Syn stimulates LD accumulation in dependence of lipid availability, restricts organelle size and ensures intracellular LD organization, which strictly depends on functional membrane-binding. Supporting a general role of Syn in cellular lipid and cholesterol metabolism, our results point to additional loss-of-function similarities between Parkinsons, Alzheimers and Gauchers disease.

cell biology↗

bioRxiv2023

The α/β hydrolase domain-containing protein 1 (ABHD1) acts as a lysolipid lipase and is involved in lipid droplet formation

Lipid droplets (LDs) are the major sites of lipid and energy homeostasis. However, few LD biogenesis proteins have been identified. Here, using Chlamydomonas as a model, we show that ABHD1, a member of the /{beta} hydrolase domain-containing protein family, is a novel type of LD-associated protein which stimulates LD formation through two distinct actions on the LD surface, one enzymatic and the other structural. ABHD1 was localized to LD surface in Chlamydomonas cells. The knockout mutants contained similar amounts of triacylglycerols (TAG) but their LDs showed an increased content in lyso- derivatives of the betaine lipid diacylglyceryl-N,N,N-trimethylhomoserine (DGTS). Over-expression of ABHD1 in Chlamydomonas induced LD formation and boosted TAG content, suggesting a key role in LD biogenesis. The purified recombinant ABHD1 protein hydrolyzed lyso-DGTS, producing a free fatty acid and a glyceryltrimethylhomoserine moiety. In vitro experiments using droplet- embedded vesicles showed that ABHD1 promoted LD emergence. Taken together, these results identify ABHD1 as a new player in LD formation by its lipase activity on lyso-DGTS and by its distinct biophysical property. This study further suggests that lipases targeted to LDs and able to act on their polar lipid coat may be interesting tools to promote LD assembly in eukaryotic cells. Significant statementLipid droplets are subcellular organelles specialized for triacylglycerol storage. Their dynamic turnover is key to managing energy homeostasis in response to cell cycle states and environmental cues. To gain insights into LD biogenesis, we characterized a putative /{beta}- hydrolase (ABHD1) in the model algae Chlamydomonas reinhardtii and show it is located at the LD surface. We found that ABHD1 overexpression promotes LD formation and acts as a lipase mainly on lyso derivatives of the betaine lipid diacylglyceryl-N,N,N-trimethylhomoserine (DGTS), the major lipid constituent of the LD hemi-membrane. We also show that ABHD1 has a remarkable biophysical property favoring LD budding. This work thus identifies a novel type of lipase acting on betaine lipid and provides a first example of a protein with a dual function nvolved in LD formation.

biochemistry↗

bioRxiv2023

Perilipin membrane association determines Lipid Droplet heterogeneity in differentiating adipocytes

The storage of fat within lipid droplets (LDs) of adipocytes is critical for whole-body health. Acute fatty acid (FA) uptake by differentiating adipocytes leads to the formation of at least two LD-classes marked by distinct perilipins (PLINs). How this LD-heterogeneity arises is an important yet unresolved cell biological problem. Here, we show that an unconventional integral membrane-segment (iMS) targets the adipocyte specific LD-surface factor PLIN1 to the endoplasmic reticulum (ER) and facilitates high affinity binding to the first LD-class. The other PLINs remain largely excluded from these LDs until FA-influx recruits them to a second LD-population. Preventing ER-targeting turns PLIN1 into a soluble, cytoplasmic LD-protein, reduces its LD-affinity and switches its LD-class specificity. Conversely, moving the iMS to PLIN2 leads to ER-insertion and formation of a separate LD-class. Our results shed light on how differences in organelle targeting and disparities in lipid-affinity of LD-surface factors contribute to formation of LD-heterogeneity. HighlightsO_LIPLIN1 is an integral membrane protein behaving as a class I LD protein. C_LIO_LIAn unconventional integral membrane segment (iMS) mediates ER insertion. C_LIO_LIThe iMS is required for LD-heterogeneity in differentiating adipocytes. C_LIO_LIHigh affinity LD-targeting from the ER is likely a gated process. C_LI eTOC blurbMajchrzak et al. use biochemistry, imaging and in vitro experiments to identify molecular features in PLIN1 that determine ER insertion and high lipid droplet affinity which are both important in the generation of LD-heterogeneity within differentiating adipocytes.

cell biology↗

bioRxiv2022

MOSPD2 a new endoplasmic reticulum-lipid droplet tether functioning in LD homeostasis

Membrane contact sites between organelles are organized by protein bridges. Among the components of these contacts, the VAP family comprises endoplasmic reticulum (ER)-anchored proteins, such as MOSPD2, functioning as major ER-organelle tethers. MOSPD2 distinguishes itself from the other members of the VAP family by the presence of a CRAL-TRIO domain. In this study, we show that MOSPD2 forms ER-LD contacts thanks to its CRAL-TRIO domain. MOSPD2 ensures the attachment of the ER to LDs through a direct protein-membrane interaction involving an amphipathic helix that has an affinity for lipid packing defects present at the surface of LDs. Remarkably, the absence of MOSPD2 markedly disturbs the assembly of lipid droplets. These data show that MOSPD2, in addition to being a general ER receptor for inter-organelle contacts, possesses an additional tethering activity and is specifically implicated in the biology of LDs via its CRAL-TRIO domain.

cell biology↗