Anderson R A, Bryson G M, Parks J S
Department of Internal Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina, 27157, USA.
Mol Genet Metab. 1999 Nov;68(3):333-45. doi: 10.1006/mgme.1999.2904.
Mechanisms producing the divergent phenotypes, Wolman disease (WD) and cholesterol ester storage disease (CESD), associated with the genetic deficiency of human lysosomal acid lipase/cholesterol ester hydrolase (hLAL) function were investigated with the determination of HLAL activity levels, mRNA and protein expression, and defects in structural gene sequences in cells from three WD and five CESD patients. Measured with natural substrates, HLAL activities were all below 2% of normal, regardless of phenotype. Immunoblotting showed a lack of detectable hLAL protein in all mutant fibroblasts. Four CESD, but no WD genomes contained at least one allele with a specific exon 8 splice junction mutation, c.894 G>A, that encodes a shortened form of hLAL mRNA. Other CESD mutations were identical in type to the WD defects: nucleotide deletions (positions 397, 684, 980), insertions (594), or substitutions (193, 347) that result in premature terminations precluding any function. The only exception was a substitution at nucleotide 866 in the CESD case without an exon 8 splicing mutation; expression of the predicted S289C change in a transfection assay produced a low, but clearly measurable, level of acid esterase activity. Although it is not easily demonstrated in conventional assays, CESD is distinct from WD in that at least one mutant allele has the potential to produce enough residual enzymatic function to ameliorate the phenotype; in the majority of CESD cases this may come from a single, easily detected, splicing mutation in one allele.
研究了与人类溶酶体酸性脂肪酶/胆固醇酯水解酶(hLAL)功能的基因缺陷相关的两种不同表型,即沃尔曼病(WD)和胆固醇酯贮积病(CESD)产生的机制,测定了3例WD患者和5例CESD患者细胞中的HLAL活性水平、mRNA和蛋白质表达以及结构基因序列缺陷。用天然底物测量时,无论表型如何,HLAL活性均低于正常水平的2%。免疫印迹显示所有突变成纤维细胞中均未检测到hLAL蛋白。4例CESD患者的基因组(但WD患者的基因组未出现这种情况)含有至少一个具有特定外显子8剪接连接突变c.894 G>A的等位基因,该突变编码一种缩短形式的hLAL mRNA。其他CESD突变在类型上与WD缺陷相同:核苷酸缺失(第397、684、980位)、插入(第594位)或替换(第193、347位),这些都会导致提前终止,从而无法产生任何功能。唯一的例外是在没有外显子8剪接突变的CESD病例中,第866位核苷酸发生了替换;在转染试验中预测的S289C变化的表达产生了低水平但明显可测量的酸性酯酶活性。尽管在传统检测中不容易证明,但CESD与WD的不同之处在于,至少一个突变等位基因有可能产生足够的残余酶功能来改善表型;在大多数CESD病例中,这可能来自一个等位基因中单个易于检测的剪接突变。