Cenedella R J
Department of Biochemistry, Kirksville College of Osteopathic Medicine, Missouri 63501, USA.
Invest Ophthalmol Vis Sci. 1995 Sep;36(10):2133-41.
To determine the principle site (epithelium or superficial cortex) of gene transcription and mRNA translation for the regulatory enzyme of lens cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR). To evaluate the contribution of waning enzyme synthesis versus enzyme turnover by proteolysis in accounting for the disappearance of HMGR protein from elongated fiber cells.
Young rats were treated with lovastatin, a drug that increases transcripts of the HMGR gene and translation of HMGR mRNA in lens secondary to inhibiting cholesterol biosynthesis. The relative concentration of HMGR mRNA in lens epithelium and superficial cortex was estimated by a competitive reverse transcriptase-polymerase chain reaction system. Relative HMGR protein levels were estimated by Western blot analysis. Because lovastatin is cleared rapidly from the lens, the half-life of HMGR protein in epithelium and cortex was estimated by following the disappearance of the increased pool of enzyme protein from each compartment with time after halting drug treatment.
Between 75% and 90% of the total content of HMGR mRNA and protein in the epithelium and the superficial cortex of control rat lens was located in the cortex. Treatment with lovastatin increased the content of the mRNA in epithelium and cortex by approximately 0.4-fold and HMGR protein content approximately 5-fold. Although the concentration of HMGR mRNA and protein was similarly increased in epithelium and superficial cortex, approximately 85% to 90% of the total increase in mRNA and protein content was located in the cortex because of that area's greater mass. The half-life for the disappearance of the increased pool of HMGR protein from epithelium and cortex was similar at approximately 14 to 17 hours.
The bulk of HMGR gene transcription and mRNA translation apparently is confined to elongating fiber cells. The 10-fold greater increase in enzyme protein than mRNA levels after lovastatin treatment indicates that enzyme concentration in lens is controlled mainly by effects on HMGR mRNA translation or rates of HMGR proteolysis. The observed rapid turnover of enzyme protein in the epithelium and the superficial cortex, if applicable to the deeper cortex and the homeostatic state (absence of drug exposure), suggests that the gradual disappearance of HMGR protein from the lens could be caused by waning of enzyme synthesis rather than to proteolysis in the absence of continuing enzyme synthesis.
确定晶状体胆固醇生物合成调节酶3-羟基-3-甲基戊二酰辅酶A还原酶（HMGR）基因转录和mRNA翻译的主要位点（上皮或浅层皮质）。评估在解释HMGR蛋白从伸长的纤维细胞中消失的过程中，酶合成减少与蛋白水解导致的酶周转的作用。
用洛伐他汀处理幼鼠，该药物通过抑制胆固醇生物合成，增加晶状体中HMGR基因的转录和HMGR mRNA的翻译。通过竞争性逆转录聚合酶链反应系统估计晶状体上皮和浅层皮质中HMGR mRNA的相对浓度。通过蛋白质印迹分析估计相对HMGR蛋白水平。由于洛伐他汀可迅速从晶状体中清除，在停止药物治疗后，通过跟踪每个区室中增加的酶蛋白池随时间的消失情况，估计上皮和皮质中HMGR蛋白的半衰期。
对照大鼠晶状体上皮和浅层皮质中HMGR mRNA和蛋白的总含量，有75%至90%位于皮质。用洛伐他汀处理使上皮和皮质中mRNA的含量增加约0.4倍，HMGR蛋白含量增加约5倍。尽管上皮和浅层皮质中HMGR mRNA和蛋白的浓度同样增加，但由于该区域质量较大，mRNA和蛋白含量总增加量的约85%至90%位于皮质。上皮和皮质中增加的HMGR蛋白池消失的半衰期相似，约为14至17小时。
HMGR基因转录和mRNA翻译的大部分显然局限于伸长的纤维细胞。洛伐他汀处理后，酶蛋白增加量比mRNA水平高10倍，表明晶状体中的酶浓度主要受对HMGR mRNA翻译的影响或HMGR蛋白水解速率的控制。上皮和浅层皮质中观察到的酶蛋白快速周转，如果适用于更深层皮质和稳态（无药物暴露），则表明晶状体中HMGR蛋白的逐渐消失可能是由于酶合成减少，而非在没有持续酶合成的情况下由蛋白水解导致。
