Meissner J D, Naumann A, Mueller W H, Scheibe R J
Zentrum Physiologie, Medizinische Hochschule Hannover, 30623 Hannover, Germany.
Biochem J. 1999 Mar 1;338 ( Pt 2)(Pt 2):561-8.
UDP-N-acetylglucosamine:dolichyl-phosphate N-acetylglucosamine-1-phosphate transferase (GPT) is the first enzyme in the dolichol pathway of protein N-glycosylation, and is implicated in the developmental programmes of a variety of eukaryotes. In the present study we describe the effects of all-trans-retinoic acid (RA) on the levels of GPT protein and enzymic activity, and on the transcription rate of the GPT gene, in mouse P19 teratocarcinoma cells. RA caused a dose-dependent and protein-synthesis-dependent induction of enzyme activity. The maximum induction of GPT activity (about 3-fold) required 2 days of exposure to 1 microM RA. Induced GPT activity also resulted in an increase in the rate of incorporation of [3H]mannose into Glc3Man9GlcNAc2. Enzymic activities paralleled GPT gene expression. The GPT gene was induced (2-fold) after 7 h of RA treatment. An approx. 3-fold increase in a 48 kDa GPT protein and approx. 4-fold increases in the levels of three GPT transcripts (1.8, 2.0 and 2.2 kb) were observed after 2 days of RA treatment. The enhanced levels of GPT protein and mRNAs began to decline 3 days after the initiation of differentiation, and GPT expression was down-regulated during cellular differentiation. GPT activity decreased about 2. 8-fold to a constant level in differentiated P19 cells. The results indicate that the RA-induced enzyme activity was mainly determined by increased transcription of the GPT gene. RA-treated P19 cells were about 4-fold more resistant to tunicamycin, a fungal antibiotic which inhibits GPT, than were control cells. In addition, GPT activity in membranes from RA-treated P19 cells exhibited approx. 4-fold increased resistance to tunicamycin compared with activity in membranes from untreated control cells, demonstrating that resistance to tunicamycin is correlated with induced GPT activity. Furthermore, increased GPT activity had regulatory significance with regard to the rate of incorporation of [3H]mannose into Glc3Man9GlcNAc2-P-P-dolichol and into glycoproteins. Together, the data provide additional insights into the hormonal regulation of GPT and present evidence that the RA-mediated induction of GPT has a regulatory impact on the dolichol pathway.
UDP-N-乙酰葡糖胺：磷酸多萜醇N-乙酰葡糖胺-1-磷酸转移酶（GPT）是蛋白质N-糖基化多萜醇途径中的首个酶，与多种真核生物的发育程序有关。在本研究中，我们描述了全反式视黄酸（RA）对小鼠P19畸胎瘤细胞中GPT蛋白水平、酶活性以及GPT基因转录速率的影响。RA引起了酶活性的剂量依赖性和蛋白质合成依赖性诱导。GPT活性的最大诱导（约3倍）需要暴露于1μM RA 2天。诱导的GPT活性还导致[3H]甘露糖掺入Glc3Man9GlcNAc2的速率增加。酶活性与GPT基因表达平行。RA处理7小时后，GPT基因被诱导（2倍）。RA处理2天后，观察到48 kDa的GPT蛋白增加约3倍，三种GPT转录本（1.8、2.0和2.2 kb）水平增加约4倍。GPT蛋白和mRNA水平的增强在分化开始3天后开始下降，并且在细胞分化过程中GPT表达被下调。在分化的P19细胞中，GPT活性下降约2.8倍至恒定水平。结果表明，RA诱导的酶活性主要由GPT基因转录增加所决定。RA处理的P19细胞对衣霉素（一种抑制GPT的真菌抗生素）的抗性比对照细胞高约4倍。此外，与未处理的对照细胞的膜中的活性相比，RA处理的P19细胞的膜中的GPT活性对衣霉素的抗性增加约4倍，表明对衣霉素的抗性与诱导的GPT活性相关。此外，增加的GPT活性对于[3H]甘露糖掺入Glc3Man9GlcNAc2-P-P-多萜醇和糖蛋白的速率具有调节意义。总之，这些数据为GPT的激素调节提供了更多见解，并提供证据表明RA介导的GPT诱导对多萜醇途径具有调节作用。
