Co-Innovation Center for Sustainable Forestry in Southern China, College of Biology and the Environment, Nanjing Forestry University, 159 Longpan Road, Nanjing, 210037, China.
Technology and Engineering Center for Space Utilization, Chinese Academy of Sciences, 9 Deng Zhuang South Rd, Beijing, 100094, China.
Appl Microbiol Biotechnol. 2018 Jan;102(1):319-332. doi: 10.1007/s00253-017-8588-7. Epub 2017 Nov 2.
The helix-loop-helix (HLH) family of transcriptional factors is a key player in a wide range of developmental processes in organisms from mammals to microbes. We previously identified the bHLH transcription factor SclR in Aspergillus oryzae and found that the loss of SclR function led to significant phenotypic changes, such as rapid protein degradation and cell lysis in dextrin-polypeptone-yeast extract liquid medium. The result implied that SclR is potentially important in both traditional fermentative manufacturing and commercial enzyme production in A. oryzae because of its effect on growth. Therefore, this study presents a comparative assessment at the proteome level of the intracellular differences between an sclR-disrupted strain and a control strain using isobaric tandem mass tag (TMT) labeling for quantification. A total of 5447 proteins were identified, and 568 were differentially expressed proteins (DEPs). Of the DEPs, 251 proteins were increased by 1.5-fold, and 317 proteins were decreased by 1.5-fold in an sclR-disrupted strain compared to the control. The comparison of the quantitative TMT results revealed that SclR was mainly involved in carbon metabolism, especially carbohydrate metabolism. In addition, an enzyme profile by a semi-quantitative method (API-ZYM) indicated that three enzymes (β-galactosidase, α-glucosidase, and α-mannosidase) were significantly less active in the ∆sclR strain than in the control. Moreover, quantitative RT-PCR showed that the expression of certain genes was changed similarly to their corresponding proteins. These results suggested that a possible function of SclR during growth of A. oryzae is its important involvement in carbohydrate metabolism.
螺旋-环-螺旋(HLH)转录因子家族是从哺乳动物到微生物等生物体中广泛发育过程的关键参与者。我们之前在米曲霉中鉴定了 bHLH 转录因子 SclR,并发现 SclR 功能的丧失导致了显著的表型变化,如在糊精-多肽-酵母提取物液体培养基中快速的蛋白质降解和细胞裂解。这一结果表明,由于其对生长的影响,SclR 在米曲霉的传统发酵生产和商业酶生产中都具有潜在的重要性。因此,本研究使用等压串联质量标签(TMT)标记进行定量,在蛋白质组水平上对 sclR 缺失菌株和对照菌株之间的细胞内差异进行了比较评估。共鉴定出 5447 种蛋白质,其中 568 种为差异表达蛋白(DEPs)。与对照相比,sclR 缺失菌株中 251 种蛋白表达增加了 1.5 倍,317 种蛋白表达减少了 1.5 倍。定量 TMT 结果的比较表明,SclR 主要参与碳代谢,特别是碳水化合物代谢。此外,半定量方法(API-ZYM)的酶谱分析表明,在 ∆sclR 菌株中,三种酶(β-半乳糖苷酶、α-葡萄糖苷酶和α-甘露糖苷酶)的活性明显低于对照。此外,定量 RT-PCR 显示某些基因的表达与其相应蛋白的变化相似。这些结果表明,SclR 在米曲霉生长过程中的一个可能功能是其在碳水化合物代谢中的重要作用。