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研究关键词：C. elegans，lipid droplets，obesity，molecular genetics

研究概述/Research

脂滴是真核生物储存脂肪的一类保守的细胞器，脂滴的大小和数量直接决定细胞、组织、个体水平上的脂肪含量，脂滴代谢紊乱可导致肥胖症、二型糖尿病、心血管疾病等代谢疾病。脂滴生成、长大、融合、水解等动态过程的核心细胞生物学原理是什么（图1）？脂滴的大小和数量是如何被脂滴衣蛋白、代谢途径、信号通路综合调控的？这些问题迄今为止仍未得到很好的解答。

Fig 1. Basic cellular processes of lipid droplets (LD, golden): de novo formation, growth, fusion, and hydrolysis. Green arrows indicate anabolic processes; red, catabolic. The linear order of these processes is hypothetical. Currently, very little is known about the morphological intermediates and molecular and cellular mechanisms of these processes.

在研究一类过氧化物酶体功能缺失的秀丽线虫突变体的基础上，我们率先鉴定了线虫脂肪存储结构是脂滴而非先前所认为的“类溶酶体”、“肠颗粒”或“不同于类溶酶体的膜泡”，厘清了线虫脂肪研究领域里概念和方法的错误并奠定了未来线虫脂滴研究的基础（Zhang et al. 2010, BMC Cell Biology）；并且，我们建立了第一例线虫脂滴调控的遗传学和细胞生物学模型和一套完整的研究手段(Zhang et al. 2010, Proc. Natl. Acad. Sci. U. S. A.)。本实验室运用这些手段，并穷尽强大的线虫分子遗传学，完成首例饱和正向遗传突变体筛选（图2），筛选到四大类超大脂滴突变体drop (lipid droplet abnormal)，这些突变体分别显着上调脂滴长大、融合等过程，或显着下调脂滴水解过程（Li et al. 2016, G3: Genes Genomes Genetics）。我们正在对这些脂滴突变模型进行分子遗传分析，并结合运用生化重建，力图深入理解这些基因编码的蛋白调控脂滴长大、融合、水解的精微原理。

Fig 2. A forward genetic screen for C. elegans mutants with supersized lipid droplets. (A) Both EMS and ENU were used to mutagenize P0 animals. F2 progeny were transferred onto NGM/OP50 plates layered with BODIPY. Candidate mutants were isolated by virtue of supersized BODIPY-positive and light-refracting globular structures visible under a fluorescence stereoscope. Progeny (>F3) of F2 isolates were then tested by postfix Oil-Red-O staining, postfix Nile Red staining, and GFP::DGAT-2 marking. Supersized lipid droplets (arrows) in drop-2(ssd14), one of the 118 mutant isolates, were readily visible in bright field and were labeled by vital BODIPY (B), by GFP::DGAT-2 (C), by postfix Nile Red with a true color of gold (D), and by postfix Oil-Red-O (E). Modified from Li et al. 2016.

本实验室当前集中研究的一个过程是脂滴融合。我们发现：CYP-37础1（细胞色素笔450蛋白家族中的一员）和贰惭叠-8（笔450氧化还原酶）共同合成一种脂溶性的激素，该激素抑制核受体顿础贵-12；当颁驰笔37-础1失去功能后，激素不能合成，顿础贵-12促进肠道细胞里的脂滴发生热敏型的融合，这种热敏型脂滴融合类似于哺乳动物褐色脂肪细胞转化为白色脂肪细胞时脂滴的变化过程（Li et al. 2017, Proc. Natl. Acad. Sci. U. S. A.）。我们正在鉴定该种激素的化学成分，并系统研究CYP-37A1—贰惭叠-8—激素—DAF-12这条信号通路调控热敏型脂滴融合的分子与细胞生物学原理（图3）。

Fig 3. Specific regulation of thermosensitive lipid droplet fusion by the CYP-37A1－贰惭叠-8－hormone－DAF-12 pathway. (A) In cyp-37A1 mutant at a high temperature of 30 degree, two lipid droplets (labeled by GFP::DGAT-2) fuse into a supersized one (arrow). The fusion is direct membrane fusion, displays no asymmetrical partnership, and finishes in less than 30 seconds. (B) The thermosensitive lipid droplet fusion is repressed in wild-type animals by the CYP-37A1－贰惭叠-8－hormone－DAF-12 pathway (colored). This pathway is different to the DAF-9－贰惭叠-8－dafachronic acids－DAF-12 pathway that regulates dauer formation, lipogenesis, and lipid droplet growth (grey). Adapted from Li et al. 2016 and Li et al. 2017.

我们还将利用线虫作为整体动物模型去筛选能正负调控脂肪贮存的化学小分子，在筛出小分子后，结合利用基因突变和体外代谢反应体系去寻找这些小分子作用的分子靶点。

我们所建立的线虫脂滴模型既能快捷地结合运用分子遗传学、细胞生物学、生物化学、脂类分析化学等技术手段，又能有效整合分子、细胞、组织、个体水平上的研究结果，这些模型能让我们迅速揭示脂滴动态细胞生物学过程的核心保守原理。因此，我们期待线虫脂滴研究将会为哺乳动物和人类脂滴研究提供可以借鉴的模式，帮助找出让人罹患脂肪代谢疾病的遗传和饮食因素，并协助预防诊断、开发新药和疗法、促进人民身体健康。

Lipid droplets are well conserved eukaryotic cellular organelles for fat storage and fat mobilization. The size and the abundance of lipid droplets directly determine the fat levels of cells, tissues, and organisms. Disorder of lipid droplet metabolism can lead to obesity, type 2 diabetes, cardiovascular disease, and other metabolic diseases. What are the core mechanisms underlying the dynamic cellular processes of lipid droplets such as de novo formation, growth, fusion, and hydrolysis (Fig. 1)? How are lipid droplet size and abundance regulated by lipid droplet coat proteins, metabolic pathways, and signaling proteins? These questions remain poorly understood to this date.

In studying a class of peroxisomal fatty acid β-oxidation mutants, my colleagues and I discovered that the cellular fat storage structures in the model organisms Caenorhabditis elegans are lipid droplets but not “lysosome-related organelles (LROs)”, “gut granules”, or the so called “vesicles distinct from LROs”. This study thus cleared off conceptual confusions and technical mistakes in the C. elegans fat research field, and, laid down a foundation for future C. elegans lipid droplet investigation (Zhang et al. 2010, BMC Cell Biology). Furthermore, we established the first genetic and cell biological model of C. elegans lipid droplet regulation and a suite of research tools (Zhang et al. 2010, Proc. Natl. Acad. Sci. U.S.A.). By exploiting these tools and exhausting the awesome power of C. elegans molecular genetics, our laboratory has conducted and reported the first saturated forward genetic screen for mutants with supersized lipid droplets (Fig. 2). These gene mutants, named as drop (lipid droplet abnormal), fall into to four classes that differentially display significant up-regulation of either lipid droplet growth or fusion, or significant down-regulation of lipid droplet hydrolysis (Li et al. 2016, G3: Genes Genomes Genetics). We are conducting molecular genetics analysis and biochemical reconstitution with these four classes of lipid droplet mutant models. We anticipate uncovering the intricate mechanisms of how proteins encoded by these genes regulate the growth, fusion, and hydrolysis of lipid droplets.

We are currently focused on the research of one of the lipid droplet processes, i.e., lipid droplet fusion. We discovered that CYP-37A1 (a member of the cytochrome P450 protein family) and EMB-8 (the P450 oxidoreductase) act together to synthesize a lipophilic hormone(s). This hormone inhibits the nuclear receptor DAF-12. When CYP-37A1 is dysfunctional, the hormone is not synthesized so that DAF-12 becomes active to promote thermosensitive lipid droplet fusion in intestine cells. The thermosensitive lipid droplet fusion is analogous to what happens to lipid droplets in the conversion of brown adipocytes to white adipocytes in mammals (Li et al. 2017, Proc. Natl. Acad. Sci. U. S. A.). We are trying to identify the chemical constituents of the putative hormone. We are also systematically probing the molecular and cellular mechanism of how the CYP-37A1－贰惭叠-8－hormone－DAF-12 pathway regulates thermosensitive lipid droplet fusion (Fig. 3).

We also like to use C. elegans as a whole animal model to screen for small chemical compounds that negatively or positively regulate fat storage and mobilization. Once we have identified these compounds, we will combine genetic mutants and in vitro metabolic assays to identify their molecular targets.

The C. elegans lipid droplet models that we have established lend us two advantages: a quick combination of molecular genetics, cell biology, biochemistry, and lipid analytical chemistry, and, an effective integration of results obtained at the levels of molecules, cells, tissues, and whole animals. These models put us on a fast track to uncovering conserved mechanisms of the dynamic processes of lipid droplets. Thus, we expect to present a C. elegans paradigm of lipid droplet regulation to benefit the research of mammalian and human lipid droplets, with an ultimate goal of identifying genetic and dietary risk factors, prevention guidelines, and therapeutic interventions for human metabolic diseases.

发表论文/Publications

1. 1. .

2. 2. . (Commented in Science China Life Sciences; reported by EurekAlert!, Mirage News, and Science X )

3. 3. .

4. 4. . (Paper downloaded the most from the journal in the month; recommended as Paper of the Month by Genetics Society of America)

5. 5. . (* co-corresponding author)

6. 6. . (Cover article; recommended by Faculty of 1000 Biology)

7. 7. . (Rated as Highly Accessed & Featured by BMC Cell Biology)

8. 8. . (Recommended by Faculty of 1000 Biology)

9. 9. Zhang SO, Mathur S, Hattem G, Tassy O, Pourquié O (2010) Sex-dimorphic gene expression and ineffective dosage compensation of Z-linked genes in gastrulating chicken embryos. BMC Genomics 11: 13.

10. 10.Zhang SO, Kuo D-H, Weisblat DA (2009) Grandparental stem cells in leech segmentation: Differences in CDC42 expression are correlated with an alternating pattern of blast cell fates. Developmental Biology 336: 112-131.

11. 11. Zhang SO, Weisblat DA (2005) Applications of mRNA injections for analyzing cell lineage and asymmetric cell divisions during segmentation in the leech Helobdella robusta. Development 132: 2103-2113.

联系方式/Contact

北京市西叁环北路105号91糖心vlog传媒

邮编100048

Shaobing O. Zhang, Professor

Capital Normal University

College of Life Sciences

#105 Xi-San-Huan-Bei-Lu

Beijing 100048, CHINA

加入我们/Join

本实验室课题组长&苍产蝉辫;2005年博士毕业于美国加州大学伯克利分校（叠别谤办别濒别测），2006-2011年先后在美国霍华德休斯医学研究所（贬贬惭滨）、斯陶尔思医学研究所（厂迟辞飞别谤蝉）从事博士后研究。自2011年在首都师大建立实验室以来，我们已利用正向遗传学筛选出一系列能正负调控线虫脂滴动态细胞生物学过程的突变体，我们也已在体外生化重建脂滴动态过程方面取得了进展。未来将综合利用正向遗传学、生化重建及其它必要的方法手段去系统地研究调控脂滴动态过程的遗传通路和核心生化原理。我们欢迎踏实勤奋的学生来本实验室研修博士学位，我们也欢迎新近的博士毕业生来本实验室从事博士后训练。课题组长着重培养学生和博士后的系统科研思维、独立动手能力，视他们的职业发展前途为本实验室最重要的目标之一，优秀的博士生将会有机会被送到国外知名高校进行交流培养、或被送出国参加学术会议。感兴趣的个人请提前、直接联系课题组长，课题组长将根据候选个人的具体情况做出录用意向。