Biomolecular Discovery Research Centre, Department of Molecular Sciences, Macquarie University, Sydney, NSW 2109, Australia.
ARC Centre for Nanoscale BioPhotonics, Department of Molecular Sciences, Macquarie University, Sydney, NSW 2109, Australia.
J Proteome Res. 2021 Feb 5;20(2):1313-1327. doi: 10.1021/acs.jproteome.0c00702. Epub 2020 Dec 31.
Human adipose tissue contains a major source of adipose-derived stem cells (ADSCs) that have the ability to differentiate into various cell types: , ADSCs can differentiate into mesenchymal lineages including adipocytes, while , ADSCs become mature adipocytes. Protein glycosylation has been shown to change in stem cell differentiation, and while ADSCs have been acknowledged for their therapeutic potential, little is known about protein glycosylation during human ADSC adipogenic differentiation. In the present study, the global membrane protein glycosylation of native adipocytes was compared to ADSCs from the same individuals as a model of adipogenesis. For adipogenesis, ADSCs were adipogenically differentiated in cell culture using an optimized, large-scale differentiation procedure. The membrane glycome of the differentiated ADSCs (dADSCs) was compared with mature adipocytes and the progenitor ADSCs. A total of 137 glycan structures were characterized across the three cell types using PGC-LC coupled with negative-ion electrospray ionization mass spectrometry (ESI-MS)/MS. Significantly higher levels of bisecting GlcNAc-type -glycans were detected in mature adipocytes (32.1% of total glycans) and in dADSC progeny (1.9% of total glycans) compared to ADSCs. This was further correlated by the mRNA expression of the MGAT3 gene responsible for the enzymatic synthesis of this structural type. The bisecting GlcNAc structures were found on the majority of human native adipocyte membrane proteins, suggesting an important role in human adipocyte biology. Core fucosylation was also significantly increased during adipogenesis but did not correlate with an increase in Fut8 gene transcript. Unexpectedly, low abundance structures carrying rare β-linked Gal-Gal termini were also detected. Overall, the -glycan profiles of the differentiated progeny did not reflect native adipocytes, and the results show that bisecting GlcNAc structures are a characteristic feature of human adipocyte membrane protein -glycosylation. Raw MS files are available on GlycoPOST (ID: GPST000153 https://glycopost.glycosmos.org/).
人类脂肪组织中含有大量脂肪来源的干细胞 (ADSCs)，这些细胞具有分化为多种细胞类型的能力：在体外培养条件下，ADSCs 可以分化为间充质谱系，包括脂肪细胞，而在体内，ADSCs 成为成熟脂肪细胞。研究表明，干细胞分化过程中蛋白糖基化会发生改变，虽然 ADSCs 的治疗潜力已得到认可，但人们对其在人类 ADSC 成脂分化过程中的蛋白糖基化知之甚少。在本研究中，以同一供体来源的原代脂肪细胞和 ADSCs 为模型，比较了其天然脂肪细胞的整体膜蛋白糖基化。为了进行成脂分化，ADSCs 在细胞培养中采用优化的大规模分化程序进行成脂分化。将分化后的 ADSCs（dADSCs）的膜聚糖组与成熟脂肪细胞和祖代 ADSCs 进行比较。使用 PGC-LC 与负离子电喷雾电离质谱（ESI-MS）/MS 相结合，共鉴定了三种细胞类型中的 137 种聚糖结构。与 ADSCs 相比，成熟脂肪细胞（总糖的 32.1%）和 dADSC 祖代（总糖的 1.9%）中检测到更高水平的双分支 GlcNAc 型聚糖。这与负责这种结构类型酶合成的 MGAT3 基因的 mRNA 表达进一步相关。双分支 GlcNAc 结构存在于大多数人源天然脂肪细胞膜蛋白上，提示其在人脂肪细胞生物学中具有重要作用。成脂分化过程中核心岩藻糖基化也显著增加，但与 Fut8 基因转录物的增加无关。出乎意料的是，还检测到低丰度带有罕见 β 连接的 Gal-Gal 末端的结构。总体而言，分化后代的 -聚糖谱并不反映天然脂肪细胞，结果表明双分支 GlcNAc 结构是人脂肪细胞膜蛋白 -糖基化的特征。原始 MS 文件可在 GlycoPOST（ID：GPST000153 https://glycopost.glycosmos.org/）上获取。
