Andrade L E, Chan E K, Raska I, Peebles C L, Roos G, Tan E M
W. M. Keck Autoimmune Disease Center, Department of Molecular and Experimental Medicine, Scripps Clinic and Research Foundation, La Jolla, California 92037.
J Exp Med. 1991 Jun 1;173(6):1407-19. doi: 10.1084/jem.173.6.1407.
Antibodies producing an unusual immunofluorescent pattern were identified in the sera of patients with diverse autoimmune features. This pattern was characterized by the presence of up to six round discrete nuclear bodies in interphase cell nuclei. Immunoblotting analysis showed that these sera recognized an 80-kD nuclear protein, and affinity-purified anti-p80 antibody from the protein band reproduced the fluorescent staining of nuclear bodies. Colloidal gold immunoelectron microscopy showed that the affinity-purified anti-p80 antibody recognized the coiled body, an ultramicroscopic nuclear structure probably first described by the Spanish cytologist Ramon y Cajal. Five cDNA clones were isolated from a MOLT-4 cell lambda gt-11 expression library using human antibody and oligonucleotide probes. The longest cDNA insert was 2.1 kb and had an open reading frame of 405 amino acids. A clone encoding a 14-kD COOH-terminal region of the protein was used for expression of a beta-galactosidase fusion protein. An epitope was present in this COOH-terminal 14-kD region, which was recognized by 18 of 20 sera with anti-p80 reactivity, and affinity-purified antibody from the recombinant protein also reacted in immunofluorescence to show specific staining of the coiled body. This is the first demonstration and molecular cloning of a protein that appears to have particular identification with the coiled body, and it was designated p80-coilin. Autoantibody to p80-coilin may be useful for the elucidation of the structure and function of the coiled body, and the availability of a cDNA sequence could be helpful in further studies to clarify the clinical significance of this autoantibody response.
在具有多种自身免疫特征的患者血清中鉴定出产生异常免疫荧光模式的抗体。这种模式的特征是在间期细胞核中存在多达六个圆形离散核体。免疫印迹分析表明，这些血清识别一种80kD的核蛋白，从蛋白带中亲和纯化的抗p80抗体重现了核体的荧光染色。胶体金免疫电子显微镜显示，亲和纯化的抗p80抗体识别卷曲小体，这是一种超微核结构，可能最早由西班牙细胞学家拉蒙·伊·卡哈尔描述。使用人抗体和寡核苷酸探针从MOLT-4细胞λgt-11表达文库中分离出五个cDNA克隆。最长的cDNA插入片段为2.1kb，有一个405个氨基酸的开放阅读框。一个编码该蛋白14kD羧基末端区域的克隆用于表达β-半乳糖苷酶融合蛋白。在这个14kD羧基末端区域存在一个表位，20份具有抗p80反应性的血清中有18份能识别该表位，从重组蛋白中亲和纯化的抗体在免疫荧光中也有反应，显示出卷曲小体的特异性染色。这是首次证明并分子克隆了一种似乎与卷曲小体有特定关联的蛋白，它被命名为p80-卷曲螺旋蛋白。针对p80-卷曲螺旋蛋白的自身抗体可能有助于阐明卷曲小体的结构和功能，cDNA序列的可用性可能有助于进一步研究以阐明这种自身抗体反应的临床意义。
