Arginine (315) is required for the PLIN2-CGI-58 interface and plays a functional role in regulating nascent LDs formation in bovine adipocytes.

Perilipin-2 (PLIN2) can anchor to lipid droplets (LDs) and play a crucial role in regulating nascent LDs formation. Bimolecular fluorescence complementation (BiFC) and flow cytometry were examined to verify the PLIN2-CGI-58 interaction efficiency in bovine adipocytes. GST-Pulldown assay was used to detect the key site arginine315 function in PLIN2-CGI-58 interaction. Experiments were also examined to research these mutations function of PLIN2 in LDs formation during adipocytes differentiation, LDs were measured after staining by BODIPY, lipogenesis-related genes were also detected. Results showed that Leucine (L371A, L311A) and glycine (G369A, G376A) mutations reduced interaction efficiencies. Serine (S367A) mutations enhanced the interaction efficiency. Arginine (R315A) mutations resulted in loss of fluorescence in the cytoplasm and disrupted the interaction with CGI-58, as verified by pulldown assay. R315W mutations resulted in a significant increase in the number of LDs compared with wild-type (WT) PLIN2 or the R315A mutations. Lipogenesis-related genes were either up- or downregulated when mutated PLIN2 interacted with CGI-58. Arginine315 in PLIN2 is required for the PLIN2-CGI-58 interface and could regulate nascent LD formation and lipogenesis. This study is the first to study amino acids on the PLIN2 interface during interaction with CGI-58 in bovine and highlight the role played by PLIN2 in the regulation of bovine adipocyte lipogenesis.

Disorder in milk proteins: adipophilin and TIP47, important constituents of the milk fat globule membrane.

Milk fat globules (MFGs), which are secreted by the epithelial cells of the lactating mammary glands, account for the most of the nutritional value of milk. They are enveloped by the milk fat globule membrane (MFGM), a complex structure consisting of three phospholipid membrane monolayers and containing various lipids. Depending on the origin of milk, specific proteins accounts for 5-70% of the MFGM mass. Proteome of MFGMs includes hundreds of proteins, with nine major components being adipophilin, butyrophilin, cluster of differentiation 36, fatty acid binding protein, lactadherin, mucin 1, mucin 15, tail-interacting protein 47 (TIP47), and xanthine oxidoreductase. Two of the MFGM components, adipophilin and TIP47, belong to the five-member perilipin family of lipid droplet proteins. Adipophilin is involved in the formation of cytoplasmic lipid droplets and secretion of MFGs. This protein is also related to the formation of other lipid droplets that exist in most cell types, playing an important role in the transport of lipids from ER to the surface of lipid droplets. TIP47 acts as a cytoplasmic sorting factor for mannose 6-phosphate receptors and is recruited to the MFGM. Therefore, both adipophilin and TIP47 are moonlighting proteins, each possessing several unrelated functions. This review focuses on the main functions and specific structural features of adipophilin and TIP47, analyzes similarities and differences of these proteins among different species, and describes these proteins in the context of other members of the perilipin family.Communicated by Ramaswamy H. Sarma.

The Expression Pattern of PLIN2 in Differentiated Adipocytes from Qinchuan Cattle Analysis of Its Protein Structure and Interaction with CGI-58.

PLIN2 (Perilipin-2) is a protein that can anchor on the membrane of lipid droplets (LDs), playing a vital role in the early formation of LDs and in the regulation of LD metabolism in many types of cells. However, little research has been conducted in cattle adipocytes. In the present study, we found that the expression of PLIN2 mRNA peaks at Day 2 during cattle adipocyte differentiation (p < 0.01), but PLIN2 protein levels maintain high abundance until Day 4 and then decrease sharply. We first built an interaction model using PyMOL. The results of a pull-down assay indicated that bovine PLIN2 and CGI-58 (ABHD5, α/β hydrolase domain-containing protein 5) had an interaction relationship. Furthermore, Bimolecular Fluorescence Complementation-Flow Cytometry (BiFC-FC) was used to explore the function of the PLIN2-CGI-58 interaction. Interestingly, we found that different combined models had different levels of fluorescence intensity; specifically, PLIN2-VN173+CGI-58-VC155 expressed in bovine adipocytes exhibited the highest level of fluorescence intensity. Our findings elucidate the PLIN2 expression pattern in cattle adipocytes, the protein structure and the function of protein⁻protein interactions (PPI) as well as highlight the characteristics of bovine PLIN2 during the early formation and accumulation of lipid droplets.

Cytoplasmic Lipid Accumulation Characteristic of the Cribriform Variant of Papillary Thyroid Carcinoma.

OBJECTIVE: The purpose of this study was to clarify the diagnostic significance of cytoplasmic lipid accumulation (CLIA) in the cribriform variant of papillary thyroid carcinoma (CV-PTC). METHODS: We performed a histological, immunohistochemical, and cytological examination of 35 CV-PTC cases at the Kuma Hospital. CLIA was defined as bubble-like multivacuolation in cytoplasm with distinct cell border. We also examined 100 conventional PTC (con-PTC) cases as controls. RESULTS: Histological analysis showed the presence of carcinoma cells with CLIA in 60.0% of CV-PTC and 5.0% of con-PTC cases. The vacuoles tended to distribute in the subnuclear portion of carcinoma cells showing papillary growth. They were positive for oil red O staining and adipophilin. The carcinoma cells without the vacuoles showed a subnuclear dot-like expression for adipophilin in CV-PTC cases, but not in the con-PTC cases. Cytological analysis showed CLIA in 17 (54.8%) of the 31 CV-PTC cases, but not in the con-PTC cases. CONCLUSION: This is the first study to report the presence of carcinoma cells with CLIA in CV-PTC. The subnuclear dot-like expression of adipophilin may be characteristic of CV-PTC. These findings might be related to degenerative changes occurring in CV-PTC.

Perilipin2 plays a positive role in adipocytes during lipolysis by escaping proteasomal degradation.

Perilipin2 (Plin2), also known as adipose differentiation-related protein (ADRP), or adipophilin, is a member of the PAT family involved in lipid droplet (LD) formation in the liver and peripheral tissues. Although Plin2 was originally identified as a highly expressed gene in adipocytes, its physiological role in mature adipocytes is largely unknown. In this report, we investigated the regulation of Plin2 expression and its function in differentiated adipocytes of mouse embryonic fibroblasts (MEFs). Plin2 mRNA levels increased during adipocyte differentiation whereas protein levels did not. Plin2 was degraded through the ubiquitin-proteasome pathway but was inhibited by lipolytic inducers. Furthermore, lentiviral-mediated Plin2 knockdown attenuated lipolysis in differentiated MEFs in a time-dependent manner. Oleic acid-induced LD formation enhanced Plin2 protein stability when it was localized to LDs. Furthermore, a mutational analysis revealed that the ubiquitination and degradation of Plin2 required both the second and third alanine in the N-terminal region. These results suggest that Plin2 is degraded in the cytosol in its N-terminal amino acid sequence-dependent manner and instead becomes stable when localized on LDs. Our findings highlight the relationship between protein stability and a previously unnoticed function of Plin2 during lipolysis in adipocytes.