飲食調節之線蟲脂肪新陳代謝

Abstract

隨著健康知識的普及，國人逐漸重視飲食裡的成分標示，食物所提供之營養素對於維持生物體內的恆定至關重要。在許多模式生物中都已證實飲食會影響到生物之生育、發育、老化等生理行為，然而其中詳細的分子機制還尚未明朗。利用ORO染色技術發現餵食Comamonas DA1877的線蟲(C. elegans) 其體內的脂肪含量比餵食標準食物E. coli OP50的線蟲還要少。因此我們利用餵食這兩種不同食物的線蟲探討飲食如何影響生物體內的脂肪代謝。我的研究中發現，線蟲體內磷脂醯膽鹼(phosphatidylcholine)的含量上升是導致DA1877餵食之三酸甘油脂(triacylglycerol)含量下降的原因之一，我們進一步發現DA1877飲食造成磷脂醯膽鹼所調控的固醇調節區域結合蛋白(SBP-1) 的脂肪生成功能的缺失。在DA1877餵食下SBP-1並沒有改變其在細胞核的表現量，因此我們認為 SBP-1進入細胞核內所需的活化過程和DA1877餵食下線蟲脂肪含量減少無關。有趣的是我們發現了SBP-1核內的轉錄輔激活因子MDT-15參與在飲食調控之脂肪含量改變。雖然mdt-15突變和sbp-1突變的線蟲在不同細菌餵食下對脂肪含量有不一樣的效果，但這可能是由於mdt-15在不同飲食下經由與不同脂肪代謝轉錄因子結合所致。我們經由檢視mdt-15的目標基因：脂肪酸去飽和酶(fatty acid desaturase): fat-5, fat-6 和fat-7發現DA1877的餵食導致fat-5 與 fat-7表現下降，但不影響fat-6。我們更進一步利用mdt-15突變證明在不同的細菌餵食下mdt-15對此兩基因展現不同的調控。此外，我們也發現相較於餵食OP50，DA1877能夠使mdt-15突變之線蟲保有更多的子代，這個現象也暗示著不同細菌餵食下的線蟲對mdt-15有著不同的需求。另外，我們發現mdt-15突變的線蟲其S-腺苷甲硫氨酸合成酶(sams-1)表現量被抑制了，而mdt-15是否透過調控sams-1來影響脂肪利用還待進一步的實驗釐清。另一方面，在冷昏迷(cold coma)實驗中發現，吃DA1877的線蟲適應寒冷環境的能力明顯下降，然而僅僅限制fat-5和fat-7表現量並不足以影響線蟲適應低溫的能力，氣相層析質譜(GC/MS)的結果顯示出DA1877餵食之線蟲體內有較少的多元不飽和脂肪酸(poly-unsaturated fatty acids)，這可能便是DA1877餵食之線蟲無法適應低溫的原因。我的研究結果顯示: 飲食會影響生物體的脂肪新陳代謝系統包含各種脂肪的組成比例，進而改變生物體的脂質恆定(lipid homeostasis)。 而mdt-15在不同飲食造成的脂肪代謝差異扮演了相當重要的角色，然而飲食如何調控mdt-15還需要我們更深入的探討。

Dietary effects on lifespan, development, fertility and other physiological processes have been shown in many model animals. Nevertheless, the underlying mechanisms remained largely unknown. In our lab, we fed Caenorhabditis elegans with an alternative bacteria Comamonas DA1877 and found that animals exhibited reduced fat storage by Oil Red O (ORO) stain compared to ones fed with the standard bacteria E. coli OP50. We performed thin-layer chromatography (TLC) analysis and found the lower level of triacylglycerol (TAG), the major lipid form for fat storage, and the increased level in phosphatidylcholine (PC), the dominant component of membrane phospholipids in DA1877-fed animals, indicating a dramatic lipid metabolism change mediated by diets. By abolishing PC synthesis, we demonstrated the increased PC level is the cause for DA1877-mediated lipid reduction. To understand how PC level regulates lipid content in DA1877-fed animals, we tested the regulation of lipogenic transcription factor SBP-1 on different diets. Loss of sbp-1 reduced the fat level in OP50-fed animals but not DA1877-fed ones. In addition, overexpressed SBP-1 restored lipid content reduction mediated by DA1877 diet. We further demonstrated the similar intensity of the intestinal nuclear GFP::SBP-1 in both DA1877- and OP50- fed animals, suggesting the constrained sbp-1-dependent lipogenesis on DA1877 is not through inhibiting sbp-1 nuclear localization. Intriguingly, loss of mdt-15, the sbp-1 co-activator, not only increased fat content but also enlarged the lipid droplet size compared to wild-type fed DA1877, while mdt-15 reduced lipid content in OP50-fed animals. Moreover, DA1877 diet resulted in decreased expression of fat-5 and fat-7 but not fat-6, all of which are mdt-15 target genes. By mdt-15 deficiency, we demonstrated that mdt-15 regulated its target genes, differentially on different diets, possibly via interaction with different transcription factors in regulation of lipid metabolism. In addition, we found mdt-15 might regulate lipid content through modulating the expression of sams-1, an important gene in methylation cycle. Last but not least, DA1877-fed worms exhibited poor ability to resist cold stress, suggesting the alteration of membrane fluidity. Gas chromatography–mass spectrometry (GC-MS) analysis revealed different diets changed worm lipid composition as DA1877-fed worms showed poor contents of polyunsaturated fatty acids (PUFAs) and more saturated fatty acids (SFAs) than OP50-fed worms. Together, our results proposed that diets affect lipid composition to regulate lipid level in C. elegans. The fat content reduction is caused by the relative high PC level in DA1877-fed animals and mdt-15 is identified to play a key role in the dietary-dependent lipid regulation mechanism. By dissecting different mdt-15 binding partners on different diets, it is anticipated to delineate how different diet affect lipid homeostasis in C. elegans.