一个Ile314Thr纯合突变所致红细胞丙酮酸激酶缺乏症家系

Analysis of a pyruvate kinase deficiency consanguineous pedigree caused by Ile314Thr homozygous mutation

目的 探讨1个丙酮酸激酶（Pyruvate kinase,PK）缺乏症家系的分子发病机制.方法 采用比色法检测该家系所有成员红细胞PK的活性;采用PCR法扩增先证者PKLR基因的12个外显子及其侧翼序列,并进行DNA测序以确定先证者突变位点;然后采用限制性内切酶酶切法对家系成员该位点进行分析.结果 先证者及其胞妹红细胞PK活性分别为5.89及3.45 U/g Hb（正常参考值：10.34～16.54 U/g Hb）,其父亲、母亲、外祖母、大姑红细胞PK活性分别为6.54、8.87、7.89、9.32 U/g Hb.先证者PKLR基因第7号外显子941 T＞C（Ile314Thr）纯合突变,并且通过cDNA序列分析,在RNA水平也得以证实.先证者妹妹同样为Ile314Thr纯合突变,先证者外祖母、父亲、母亲、大姑均为Ile314Thr杂合突变.结论 Ile314Thr纯合突变是该家系PK缺乏症的分子发病机制.

Objective To screen potential mutation and explore the underlying mechanism for a consanguineous pedigree featuring pyruvate kinase （PK） deficiency.Methods The red blood cell pyruvate kinase activities of all family members were detected.All the exons and intron-exon boundaries of the PKLR gene for the proband were amplified and analyzed by direct sequencing.Restriction endonuclease enzymes were used to identify the presence of mutations of all family members.Results The pyruvate kinase activities were 5.89 U/g Hb in the proband,3.45,6.54,8.87,7.89,9.32 U/g Hb in his younger sister,father,mother,grandmother and elder aunt,respectively.The homozygous missense mutation of T＞C transition at position 941 in exon 7 of PKLR gene resulted to a Ile314Thr substitution in the proband,and mutant alleles were identified at the level of RNA transcript by cDNA sequence analysis.His younger sister was also homozygous for Ile314Thr.Heterozygosity for Ile314Thr was confirmed in his grandmother,parents and elder aunt.Conclusions Ile314Thr homozygous missense mutation in exon 7 of PKLR is the molecular mechanism of pyruvate kinase deficiency in this family.