Objective

Characterization of a deletion in the exon 1 and 5′ regulatory region of the GHRHR gene in a proband with isolated growth hormone deficiency.

Methods

Multiple ligation dependent probe amplification (MLPA) assay was carried out to confirm the homozygous deletion which was suspected during screening of the GHRHR gene by single strand conformation polymorphism. A series of short range PCR amplifications were carried out to map the approximate location of the break points of the deletion. Sanger sequencing was carried out to locate the break points and to identify the length of the deletion. Long range PCR amplification was carried out to confirm the length of the deletion and to screen the parents of the proband for the deletion.

Results

A homozygous deletion was confirmed via MLPA assay. Zones of sequence similarity between upstream intergenic region and intron 1 of the GHRHR gene were identified. Break points of the deletion were identified within perfectly matching 32 bp repeat sequences ie: microhomologies in the specified zones. The novel deletion may have arisen via Alu specific microhomology mediated non-recurrent rearrangement in the maternal lineage of the proband. The deletion being reported in this study include, last 3118 bp from the upstream intergenic region and complete exon 1 and first 2620 bp from intron 1 and one of the 32 bp microhomologies. The total length of the deleted segment was 5875 bp. As the deleted region contained significant elements essential for gene expression, the identified deletion is being reported as likely pathogenic. The same deletion was identified in the mother in heterozygous state.

Conclusion

We have characterized a novel deletion that seems to have arisen via Alu specific microhomology mediated non-recurrent rearrangement at GHRHR gene locus. HGVS nomenclature of the deletion is c.-3166_58-2057del. This novel structural variant was identified to be the cause of IGHD of the affected proband.

中文翻译：

---

在GHRHR基因位点的新型总体缺失可能是由在患有孤立的生长激素缺乏症的斯里兰卡患者中发现的Alu特异性微同源性介导的。

目的

GHRHR基因外显子1和5'调控区中缺失的特征，该患者患有孤立的生长激素缺乏症。

方法

进行了多次连接依赖性探针扩增（MLPA）测定，以确认纯合缺失，该缺失在通过单链构象多态性筛选GHRHR基因时被怀疑。进行了一系列的短程PCR扩增，以定位缺失的断裂点的大致位置。进行了桑格测序以定位断裂点并鉴定缺失的长度。进行了远距离PCR扩增，以确认缺失的长度并筛选先证者的父母是否缺失。

结果

通过MLPA分析证实纯合的缺失。确定了上游基因间区域与GHRHR基因的内含子1之间的序列相似性区域。在完全匹配的32 bp重复序列（即指定区域中的微同源性）内确定了缺失的断裂点。新型删除可能是通过Alu引起的先证者的母系中特定的微同源性介导的非复发性重排。该研究报告的缺失包括：上游基因间区域的最后3118 bp和完整的外显子1，内含子1的第一个2620 bp，以及32 bp的微观同源性之一。缺失片段的总长度为5875bp。由于缺失的区域包含基因表达必不可少的重要元素，因此已报告鉴定出的缺失可能是致病的。在杂合状态的母亲中鉴定出相同的缺失。

结论

我们已经表征了一种新型的缺失，似乎是通过Alu特异性微同源性介导的GHRHR基因位点的非复发性重排而引起的。缺失的HGVS命名法是c.-3166_58-2057del。该新颖的结构变异被确定为受影响先证者IGHD的病因。