Identification of novel OCRL isoforms associated with phenotypic differences between Dent disease-2 and Lowe syndrome
Identification of novel OCRL isoforms associated with phenotypic differences between Dent disease-2 and Lowe syndrome
复制标题
鉴定与 Dent 病 2 型和 Lowe 综合征表型差异相关的新型 OCRL 亚型
DOI：
10.1093/ndt/gfab274
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发表时间：
2021
影响因子：
6.1
通讯作者：
Horinouchi Tomoko
中科院分区：
文献类型：
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作者：
Sakakibara Nana;Ijuin Takeshi;Horinouchi Tomoko
虽然Lowe综合征和Dent病-2是由Oculocerebrorenal综合征of Lowe(OCRL)突变引起的，但它们的临床严重程度有很大的不同，其分子机制尚不清楚。外显子1-7的截短突变导致Dent病-2，而外显子8-24的截断突变导致Lowe综合征。在此，我们确定了新的OCRL蛋白异构体的作用机制。方法从健康对照组和Dent病-2患者的尿液来源的培养细胞中提取信使RNA样本，检测OCRL异构体的5‘末端。结果成功地克隆了OCRL6-24外显子的新的异构体转录本，包括外显子8的翻译起始密码子。在体外蛋白表达分析中，检测到两种不同大小的蛋白质(105和80 kDa)从全长OCRL中翻译而来，而从异构体和Dent病-2变异体中只发现了一个蛋白质(80 KDa)。没有观察到Lowe综合征变异体的蛋白表达。同工酶活性与全长OCRL相当；Dent病-2变异体保持50%的酶活性，而Lowe综合征变异体保持20%的酶活性。结论我们阐明了OCRL相关疾病两种不同表型的分子机制，从外显子8开始翻译的功能性OCRL异构体与这一机制有关。
BackgroundAlthough Lowe syndrome and Dent disease-2 are caused byOculocerebrorenal syndrome of Lowe(OCRL) mutations, their clinical severities differ substantially and their molecular mechanisms remain unclear. Truncating mutations inOCRLexons 1–7 lead to Dent disease-2, whereas those in exons 8–24 lead to Lowe syndrome. Herein we identified the mechanism underlying the action of novel OCRL protein isoforms.MethodsMessenger RNA samples extracted from cultured urine-derived cells from a healthy control and a Dent disease-2 patient were examined to detect the 5′ end of theOCRLisoform. For protein expression and functional analysis, vectors containing the full-lengthOCRLtranscripts, the isoform transcripts and transcripts with truncating mutations detected in Lowe syndrome and Dent disease-2 patients were transfected into HeLa cells.ResultsWe successfully cloned the novel isoform transcripts fromOCRLexons 6–24, including the translation-initiation codons present in exon 8.In vitroprotein-expression analysis detected proteins of two different sizes (105 and 80 kDa) translated from full-lengthOCRL, whereas only one protein (80 kDa) was found from the isoform and Dent disease-2 variants. No protein expression was observed for the Lowe syndrome variants. The isoform enzyme activity was equivalent to that of full-length OCRL; the Dent disease-2 variants retained >50% enzyme activity, whereas the Lowe syndrome variants retained <20% activity.ConclusionsWe elucidated the molecular mechanism underlying the two different phenotypes inOCRL-related diseases; the functional OCRL isoform translated starting at exon 8 was associated with this mechanism.