应用全外显子测序技术筛查LEOPARD综合征患者致病基因一例

目的:报道一例LEOPARD综合征(LS)患者并检测其基因突变。方法:收集患者临床资料，提取该患者及父亲外周血DNA,应用全外显子组高通量测序检测技术，筛查患者致病基因。结果:全外显子测序结果发现患者PTPN11基因12号外显子第1415位核苷酸发生c.1415C>T杂合突变，导致第472号氨基酸由Thr变为Met(p.Thr472Met)。结论:PTPN11基因12号外显子c.1415C>T (p.Thr472Met)突变为该例LEOPARD综合征的致病突变位点。     

一例Bjornstad综合征患儿BCS1L基因突变检测

目的：检测Bjornstad综合征患儿的基因突变。方法：提取患儿，其表型正常父母及100名正常人的外周血DNA，采用二代皮肤靶向测序包检测患儿的基因突变，然后应用Sanger测序方法进行验证。结果：测序结果发现患儿及其父母在BCS1L基因存在2个杂合突变，在第7个外显子上发现c.818delC缺失突变，在第8个外显子上发现c.917G>A错义突变。健康对照中未检测到BCS1L基因突变。结论：该患儿存在BCS1L基因突变，可能与Bjornstad综合征发病有关。     

Carvajal综合征一例伴桥粒斑蛋白基因新突变

【摘要】 目的 对1例临床表现为羊毛状发，膝盖、掌跖角化性皮损，暂无心脏症状的患儿进行基因突变检测。方法 收集患儿及其父母的临床资料。提取患儿、其父母及100例无关健康对照者外周血DNA，采用二代皮肤靶向测序包检测患儿的基因突变，应用Sanger测序法进行验证。结果 患儿女，3岁，出生头发卷曲，8月龄出现掌跖角化并渐累及膝盖，其父母表型正常。测序发现，患儿桥粒斑蛋白（DSP）基因第23号外显子存在移码突变c.5152dupT（p.L1718Ffs*15），DSP基因第24号外显子检测到无义突变c.C6478T（p.R2160X）。其母亲DSP基因第23号外显子亦存在c.5152dupT移码突变，但第24号外显子未检测到相关突变。其父亲及100例健康对照中均未检测到相关突变。诊断：Carvajal综合征。结论 该例Carvajal综合征患儿存在DSP基因复合杂合突变c.5152dupT（p.L1718Ffs*15）和c.C6478T（p.R2160X），可能与其发病有关。     

双下肢浅表播散型汗孔角化症一例基因突变检测

目的：检测一例局限于双下肢的散发浅表播散型汗孔角化症患者致病基因突变。方法：提取患者外周血DNA,在Illumina HiSeq测序平台进行全基因组外显子测序,将测序结果与既往报道的汗孔角化症致病基因比对,采用Sanger测序对发现的突变位点在100例患者中进行验证。结果：最终将DSP的致病基因锁定为位于12号染色体的MVK基因,检测到10号内含子c.1040-2A>C突变位点,在全部正常对照中未检测到该位点。经检索,该突变为浅表播散型和播散性浅表性光线型汗孔角化症的共同突变位点。结论：本家系中MVK基因突变位点(c.1040-2A>C)与浅表播散型汗孔角化症发病相关。     

慢性家族性良性天疱疮五家系ATP2C1基因突变分析

目的：检测中国汉族慢性家族性良性天疱疮5家系ATP2C1基因突变情况。方法：提取5家系中12例患者、13名表型正常及100名正常对照外周血基因组DNA，经PCR扩增后进行DNA测序，并使用Chromas软件解析。结果：家系1中3例患者存在ATP2C1基因第18号外显子c.1738A>G（p.I580V）突变，家系2 中2例患者存在ATP2C1基因第25号外显子c.2416C>T(p.R806*)突变，家系3中3例患者存在ATP2C1基因第15号外显子c.1250G>A（p.R417K）突变，家系4和家系5 中ATP2C1基因未发现突变。上述家系内表型正常个体及100名正常对照中均未检测到相应突变。结论：ATP2C1基因突变可能在3例汉族HHD家系内发挥致病作用。     

有汗型外胚叶发育不良患者GJB6基因突变检测

目的:检测以脱发、甲营养不良为主要表现的有汗型外胚叶发育不良一家系的GJB6基因突变情况。方法:收集患者临床资料,提取患者及其相关亲属外周血DNA,采用PCR扩增GJB6基因编码区的全部外显子及其侧翼序列并测序。结果:基因检测发现患者GJB6基因编码序列第263号核苷酸发生c.263CT(p.A88V)杂合突变,突变来自于父亲。结论:GJB6基因编码序列第263号核苷酸发生c.263CT(p.A88V)杂合突变可能为引起该患儿及患病亲属临床表现的原因。     

遗传性对称性色素异常症一家系ADAR基因检测

目的：检测遗传性对称性色素异常症一家系的ADAR基因突变。方法：提取家系中患者、健康成员及无血缘健康对照人群外周血样DNA,PCR扩增ADAR基因外显子后测序。结果：该家系中患者均存在ADAR基因第2号外显子,第982位碱基突变（c.982C>T,p.R328X）,突变导致第328位的精氨酸被终止密码替代。家系中健康成员及健康对照人群未发现该突变。结论：该遗传性对称性色素异常症家系患者中的ADAR基因突变（c.982C>T,p.R328X）可能与发病有关。     

毛囊角化病一家系及三例散发患者ATP2A2基因突变研究

目的：对毛囊角化病（Darier's disease, DD）一家系及三例散发患者进行ATP2A2基因的突变分析。方法：收集先证者及其家系成员、散发病例的临床资料和外周血,采用PCR技术扩增ATP2A2基因所有编码区及侧翼序列,用Sanger法测序检测潜在的突变,选取与患者无亲缘关系的100例健康人作为对照,同时对已报道的ATP2A2基因突变进行文献回顾。结果：家系中三例患者均检测出ATP2A2基因第5号外显子c.380 G>A(p.G127D)新发错义突变;散发患者S1检测出第13号外显子C.1676G>A(p.R559Q)错义突变,散发患者S2检测出第14号外显子c. 2001C>T(p.D667D)同义突变,散发患者S3未检测出突变。结论：本研究中共发现三个突变,其中c.380G>A(p.G127D)在中国人群中首次报道,拓展了ATP2A2的基因突变谱。     

KRT5基因新生突变致重度型单纯型大疱性表皮松解症一家系

【摘要】 目的 报道1例重度型单纯型大疱性表皮松解症,并检测其基因突变。方法 收集患者及其父母资料和外周血,提取基因组DNA,全外显子组测序筛查患儿致病基因,随后采用Sanger测序对家系成员进行验证。结果 患者KRT5基因第7号外显子第1 429位碱基发生G→A（c.1429G>A）杂合突变,导致KRT5基因所编码的蛋白第477位谷氨酸转换成赖氨酸（p.Glu477Lys）,其父母未发现该突变。结论 该例重度型单纯型大疱性表皮松解症患者存在KRT5基因c.1429G>A（p.Glu477Lys）致病突变,属新生突变。     

单纯型大疱性表皮松解症患儿的角蛋白5基因突变研究

目的：对1例单纯型大疱性表皮松解症(EBS)患儿及其家系进行基因突变检测和致病性分析。方法：应用高通量二代测序技术捕获目标序列,对患儿进行全外显子组测序。发现致病位点后,应用Sanger测序法进行家系验证,查阅人类基因突变数据库(HGMD),运用生物信息学蛋白功能预测软件,分析变异位点的致病性。结果：患儿角蛋白5(KRT5)基因的第一外显子检出杂合变异c.536T>C(p.F179S),该变异造成KRT5蛋白的第179位氨基酸改变,患儿父母均未检测到相同突变。结论：患儿KRT5基因的杂合变异c.536T>C(p.F179S)为新发致病性变异,导致患儿KRT5缺陷,进而引发疱疹样型EBS(DM-EBS)。该变异位点在国内未见报道,扩大了我国人群KRT5的基因突变谱,为家系的遗传咨询和产前诊断提供依据。     

Nonsense mutations in the hairless gene underlie APL in five families of Pakistani origin

BACKGROUND: Atrichia with papular lesions (APL) is a rare autosomal recessive form of inherited alopecia. Affected individuals present with a distinct pattern of total hair loss on the scalp, axilla and body shortly after birth and are essentially devoid of eyelashes and eyebrows. This form of hair loss is irreversible and the histology is consistent with an absence of mature hair follicles. In addition to total atrichia, APL patients also present with papules and follicular cysts filled with cornified material. Mutations in the Hairless (HR) gene have been shown to underlie APL. OBJECTIVE: Here, we studied five unrelated large Pakistani families with clinical manifestations of APL. METHODS: Based on previous reports of HR mutations in APL, we performed direct DNA sequencing analysis. RESULTS: DNA sequencing of the HR gene in APL patients revealed three novel nonsense mutations in five unrelated families. All affected individuals were homozygous for a nonsense mutation due to C-to-T transitions at different positions in the amino acid sequence. Two families carry the mutation Q323X (CAG-TAG) in exon 3, two families harbor the mutation Q502X (CAG-TAG) in exon 6, and one family had a mutation at R940X (CGA-TGA) in exon 14. Haplotype analysis revealed that all affected individuals of both APL1 and APL16 families were homozygous for the same haplotype, and likewise, the mutation in families APL2 and APL19 was on the same haplotype. CONCLUSIONS: We report three novel nonsense mutations in the HR gene in APL. Two of the newly identified mutations, Q323X and Q502X, were found to be shared between unrelated families and marker analysis confirmed an identical homozygous haplotype for APL1 and APL16, and for APL2 and APL19. These findings suggest that Q323X and Q502X did not arise independently, but instead appear to have been propagated in the population. Collectively, these findings contribute further evidence for the involvement of hairless mutations in papular atrichia.     

Mutations in the hairless gene underlie APL in three families of Pakistani origin

BACKGROUND: Atrichia with papular lesions (APL) (OMIM#209500) is a rare autosomal recessively inherited form of irreversible alopecia characterized by papular lesions of keratin-filled cysts on various regions of the body. Males and females are equally affected and present with a distinct pattern of total hair loss on scalp, axilla and body. It begins shortly after birth with the development of hair loss, and patients are normally devoid of eyelashes and eyebrows. Mutations in the hairless (HR) gene have been previously shown to be responsible for APL. OBJECTIVE: In this study, we studied the molecular basis of APL in three unrelated families of Pakistani origin. METHOD: Molecular analysis of the HR genes was performed on genomic DNA from probands and family members. RESULTS: DNA sequencing of the HR gene in family A revealed a novel homozygous 2bp deletion in exon 6 leading to a frameshift and a downstream premature termination codon in exon 8 (1782-83delAG). In family B, we identified a novel homozygous deletion of a G nucleotide at the exon 15-intron 15 boundary, termed 3097delG. Family C carries a previously reported missense mutation consisting of an A-to-G transition at nucleotide 276 resulting in the mutation N970S in exon 14. CONCLUSION: Two mutations identified in this study are novel mutations in the HR gene and extend the body of evidence implicating the hairless gene family in the pathogenesis of human skin disorders. The one previously reported mutation suggests it may represent a recurrent mutation, or alternatively, an allele that is widely dispersed around the world.     

Identification of a novel splice site mutation in the human hairless gene underlying atrichia with papular lesions

Atrichia with Papular Lesions (APL) is a rare autosomal recessive disorder characterized by complete hair loss that begins shortly after birth with the development of papular lesions on various regions of the body. Since the establishment of hairless (HR) gene mutations as the cause of this disorder, several patients previously assumed to suffer from alopecia universalis have been subsequently diagnosed with APL. In this study we have identified a novel splicing mutation, IVS8+2T-->G, in the hairless gene. This mutation most likely abolishes normal splicing of exon 8 and potentially leads to out-of-frame skipping of this exon and a downstream premature termination codon (PTC). Our findings contribute to the growing body of HR mutations implicated in APL and provide further evidence for the differentiation of APL from alopecia universalis.     

Novel compound heterozygous nonsense mutations in the hairless gene causing atrichia with papular lesions

BACKGROUND: Atrichia with papular lesions (APL) is a rare autosomal recessive condition resulting from mutations in the hairless (HR) gene. OBJECTIVE: In the present study, we investigated the molecular basis of APL in a non-consanguineous Korean family. METHODS: Direct automated DNA sequencing of the HR gene and restriction digestion analysis were used to identify and confirm the mutation in our proband. RESULTS: Sequencing of the HR gene revealed two novel nonsense mutations in exons 2 and 4 which were subsequently confirmed via enzymatic restriction. No mutations have previously been detected in this population. CONCLUSION: The growing number of heterozygous mutations in non-consanguineous pedigrees supports the hypothesis that APL is more common than previously expected.     

Identification of a recurrent mutation in the human hairless gene underlying atrichia with papular lesions

Identification of mutations in the hairless (HR) gene in patients with atrichia with papular lesions (APL) has proven of critical importance, as it provides a basis for the differentiation between APL and alopecia universalis. The establishment of the diagnostic criteria for APL has triggered the identification of a large number of APL patients among those suspected to suffer from alopecia universalis. This advancement has resulted in the discovery of an increasing number of hairless mutations in both consanguineous and nonconsanguineous APL families. Here, we report the identification of a homozygous mutation, 3434delC, in an APL patient of Arab-Palestinian descent. The proband is a 23-year-old female with generalized scalp and body alopecia. To confirm the diagnosis of APL and to identify the specific mutation, we sequenced the hairless gene. Sequencing of all exons of the hairless gene revealed a homozygous frameshift mutation, 3434delC, in exon 18. Interestingly, the same mutation was previously identified in an Arab-Israeli family. Our data suggest that the 3434delC mutation most likely represents a founder mutation in this geographical region.     

一例伴丘疹性损害的先天性无毛症患者的基因突变研究

目的 研究1例伴丘疹性损害的先天性无毛症患者基因突变情况.方法 采用PCR和DNA直接测序法检测伴丘疹性损害的先天性无毛症HR基因的突变.同时对2个与秃发相关的基因GJB6和CDSN基因进行突变检测.结果 在HR基因、GJB6基因和CDSN基因所检测的区域中均未检测到突变.在HR基因和CDSN基因上发现了数处单核苷酸多态性(SNP).结论 在该伴丘疹性损害的先天性无毛症家庭中未检测到HR、GJB6和CDSN基因的突变.     

A newly identified missense mutation of the HR gene is associated with a novel, unusual phenotype of Marie Unna Hereditary Hypotrichosis 1 including limb deformities

Marie Unna Hereditary Hypotrichosis 1 (MUHH1; OMIM 146550), a rare monogenic condition characterized by the development of sparse, twisted hair or complete hair loss, is the consequence of mutations located in the hairless (HR) gene. We have identified a 68-year-old Hungarian woman affected by alopecia universalis and limb deformities of all four extremities. Direct sequencing of the coding regions of the HR gene revealed a novel missense mutation in the third exon of the HR gene (c.974G/A, p.Gly325Asp). The affected family member carried the mutation in a heterozygous form, while the only available, clinically unaffected family member (the son of the patient) and the unrelated controls carried the wild type sequence. The association between the presence of HR gene mutations and the development of alopecia is well-established, however, further studies are needed to elucidate the putative role of this novel HR mutation in the development of limb deformities.     

Identification of mutations in the human <Emphasis Type="Italic">hairless</Emphasis> gene in two new families with congenital atrichia

Congenital atrichia (AUC) is a form of isolated alopecia with an autosomal recessive mode of inheritance. Patients are born with normal hair but this is shed almost completely during the first weeks or months of life and never regrows. In many families the development of papular lesions is noted as an additional phenotypic feature, which defines a related phenotype designated as atrichia with papular lesions (APL). Using positional cloning strategies and the molecular findings in hairless recessive (hr/hr) mice, an animal model for AUC, mutations in the human hairless gene (HR) have been identified as a cause of AUC and APL. To date, more than 20 different mutations of the HR gene have been reported in AUC and APL including different mutation types scattered over the entire HR gene length. In this report, we describe two families of Saudi Arabian and Jewish Iranian origin comprising a number of individuals with clinical features suggestive of AUC. We therefore hypothesized that affected members may carry mutations in the HR gene. After sequencing the complete coding region of the HR gene in the Saudi Arabian family, we identified a homozygous insertion of a G (c.2661dupG; p.Thr888DfsX38) in exon 12, resulting in a premature stop codon. In a Jewish Iranian patient, we identified a homozygous splice site mutation c.1557-1G > T in intron 4. The latter mutation has been previously reported in a compound heterozygous state. In the present report, we describe the second exonic insertion mutation in the human HR gene and the first mutation in exon 12. Our study emphasizes the importance of sequencing the complete coding sequence and exon/intron junctions in the molecular diagnostics of AUC and APL.