Function of the gene of Wiskott-Aldrich syndrome

1) Wiskott-Aldrich综合征（WAS）中WASP基因突变5例WAS中发现WASP基因突变。病例1和病例2外显子1有1个碱基缺失(G)，病例3外显子2有TCA*TAA变化，病例4外显子10有CGA*TGA变化，病例5外显子10有1个碱基缺失(C)。这些突变分别导致63、63、53,320和444个氨基酸的WASP被截断。2)病例1和病例2采用PCR- sscp法检测家族成员中wascarrier的携带状态，病例4采用Xhol酶切法检测，病例3采用PCR产物直接测序法检测。5.3)采用Northern blot法检测健康志愿者单核细胞和粒细胞中WASPmRNAWASPmRNA的表达。未检出WASPmRNA。在任何来源的人类白血病细胞系中都发现了mRNA，但在人类成纤维细胞中没有发现。4) WASP多克隆抗体和单克隆抗体的制备及Western blot检测。制备了针对WASP多肽的多克隆抗体和单克隆（5A5）抗体，证明其对WASP具有特异性。利用这些抗体发现WASP在患者的单核细胞中不表达。5)用5A5单克隆抗体WASP流式细胞术检测患者淋巴母细胞样细胞系细胞质中WASP与正常志愿者相比明显阴性。然而，强非特异性结合干扰了患者单核细胞、淋巴细胞和粒细胞中WASP的检测。6) WAS患者行骨髓移植并发EBV相关B淋巴细胞增生性疾病1例，经供体淋巴细胞输注治疗成功。

1) Mutation of WASP gene in Wiskott-Aldrich syndrome (WAS)Mutation of WASP gene was found in 5 cases of WAS.One base (G) deletion in the exon 1 in cases 1 and 2, TCA*TAA change in the exon 2 in case 3, CGA*TGA change in the exon 10 in case 4 and one base deletion (C) in the exon 10 in case 5 were found. These mutations were suggested to give rise to truncated WASP of 63,63,53,320 and 444 amino acids, respectively.2) Detection of carrier status of WASCarriers in family members were detected by PCR-SSCP in case 1 and 2, by Xhol digestion in case 4 and by direct sequencing of PCR product in case 5.3) Expression of WASPmRNAWASPmRNA was expressed in mononuclear cells and granulocytes from healthy volunteers by Northern blot analysis. WASPmRNA was not detected in those from the patients. The mRNA was found in human leukemia sell lines in any origin, but not in human fibroblasts.4) Production of polyclonal and monoclonal antibodies to WASP and detection of WASP by Western blot analysisPolyclonal and monoclonal (5A5) antibodies were made to WASP peptides and proved to be specific to WASP.Using these antibodies WASP was shown not to express in mononuclear cells of the patients.5) Flow cytometric detection of cytoplasmic WASP using the 5A5 monoclonal antibodyWASP was clearly negative in cytoplasm of the lymphoblastoid cell lines from the patients as compared in that from normal volunteers. However, strong nonspecific binding interfered the detection of WASP in monocytes, lymphocytes and granulocytes from the patients.6) Case report of WAS who received bone marrow transplantation and subsequent EBV related B lymphoproliferative diseaseThe patient was treated by donor lymphocyte transfusion successfully.