Impairment of MHC class I transcription in a mutant bovine B cell line.
Impairment of MHC class I transcription in a mutant bovine B cell line.
复制标题
突变牛 B 细胞系 MHC I 类转录受损。
作者：
J. Harms;G. Splitter
为了更好地确定牛主要组织相容性复合体(MHC)抗原的调节和表达，对牛B淋巴母细胞系BL3进行了伽玛射线照射，并对存活细胞进行了MHC I类抗原丢失的免疫筛选。由此产生的I类表达缺失变异体BL3.1，在蛋白质和遗传水平上进行了鉴定，以确定缺陷的性质。显微荧光分析显示，BL3.1细胞上所有I类产物的表面密度比亲本BL3细胞降低了3-5倍。这种表面表达的降低是MHC I类特异性的，而不是MHC II类或非MHC相关基因产物免疫球蛋白(Ig)的。Northern和定量狭缝杂交分析表明，BL3.1中的I类RNA相应减少，表明存在转录水平的缺陷。核径流和转录抑制实验证实没有转录后的变化，而Southern印迹分析没有提供I类基因内部或附近发生变化的证据。为了帮助阐明I类基因表达改变的机制，将亲本BL3和变异体BL3.1与已知促进MHC I类转录的因子一起培养。干扰素、脂多糖和活化的外周血淋巴细胞培养上清液诱导的MHC-I类转录和表面表达比未经处理的对照组高2-3倍。因此，很可能在BL3.1内发生了I类基因之外的基因改变，从而损害了MHC I类的表达。
To better define the regulation and expression of bovine major histocompatibility complex (MHC) antigens, the bovine B lymphoblastoid cell line, BL3, was exposed to gamma-irradiation and surviving cells were immunoselected for MHC class I antigen loss. The resulting class I expression loss variant, BL3.1, was characterized at both the protein and genetic levels to ascertain the nature of the defect. Microfluorimetry analysis revealed a 3--5-fold surface density reduction of all class I products on BL3.1 cells relative to the parental BL3 cells. This decreased surface expression was specific for MHC class I and not for MHC class II or the non-MHC-linked gene product, immunoglobulin (Ig). Northern and quantitative slot blot analyses demonstrated a corresponding diminution of class I RNA in BL3.1 suggesting a transcriptional level defect. Nuclear run-off and transcription inhibition experiments confirmed no post-transcriptional changes while Southern blot analysis provided no evidence for alterations within or near the class I genes. To help elucidate the mechanism of altered class I expression, the parent, BL3, and variant, BL3.1, were cultured with factors known to enhance MHC class I transcription. Interferon (IFN)-gamma, lipopolysaccharide (LPS), and activated peripheral blood lymphocyte (PBL) supernatant cultured with both cell lines induced MHC class I transcription and surface expression 2--3-fold greater than the untreated controls. It is likely, therefore, that a genetic alteration outside of the class I genes has occurred within BL3.1 impairing expression of MHC class I.