Akiyama J, Volik S V, Plajzer-Frick I, Prince A, Sago H, Weier H U, Vanderbilt J N, Hawgood S, Poulain F R
Cardiovascular Research Institute and Department of Pediatrics, University of California San Francisco, San Francisco, USA.
Am J Respir Cell Mol Biol. 1999 Aug;21(2):193-9. doi: 10.1165/ajrcmb.21.2.3681.
Three of the four known mouse collectin genes have been mapped to chromosome 14. To further characterize the spatial relationship of these genes, a bacterial artificial chromosome (BAC) library of mouse chromosome 14 was screened using mouse surfactant protein (SP)-A and -D complementary DNAs (cDNAs). One large clone hybridized to both SP-A and SP-D cDNAs and was found by polymerase chain reaction (PCR) to contain sequences from one of the mouse mannose-binding lectin genes (Mbl1). We used Southern mapping and subcloning of overlapping restriction fragments to characterize the gene locus. Mapping was confirmed by fluorescent in situ hybridization of fiber-stretched BAC DNA and by Southern hybridization of restriction endonuclease-digested and PCR-amplified genomic DNA. We found that the SP-A, Mbl1, and SP-D genes reside contiguously within a 55-kb region. The SP-A and Mbl1 genes are in the same 5' to 3' orientation and 16 kb apart. The SP-D gene is in the opposite orientation to the two other collectin genes, 13 kb away from the 3' end of the Mbl1 gene. The mouse SP-D gene had not previously been characterized. We found its size (13 kb) and organization to be similar to that of human SP-D. Exon I is untranslated. The second exon is a hybrid exon that contains signal for initiation of translation, signal peptide, N-terminal domain, and the first seven collagen triplets of the collagen-like domain of the protein. Four short exons (III through VI) encode the collagen-like domain of the protein, and exons VII and VIII the linking and the carbohydrate-recognition domains, respectively.
已知的4个小鼠凝集素基因中的3个已被定位到14号染色体上。为了进一步阐明这些基因的空间关系,利用小鼠表面活性蛋白(SP)-A和-D互补DNA(cDNA)筛选了小鼠14号染色体的细菌人工染色体(BAC)文库。一个大的克隆与SP-A和SP-D cDNA均杂交,通过聚合酶链反应(PCR)发现其包含一个小鼠甘露糖结合凝集素基因(Mbl1)的序列。我们采用Southern定位和重叠限制片段亚克隆来鉴定该基因位点。通过纤维拉伸BAC DNA的荧光原位杂交以及限制内切酶消化和PCR扩增的基因组DNA的Southern杂交证实了定位。我们发现SP-A、Mbl1和SP-D基因在一个55 kb的区域内连续排列。SP-A和Mbl1基因的方向为5'至3'相同,相距16 kb。SP-D基因与其他两个凝集素基因方向相反,距离Mbl1基因的3'端13 kb。小鼠SP-D基因此前未被鉴定。我们发现其大小(13 kb)和结构与人SP-D相似。外显子I是非翻译的。第二个外显子是一个杂合外显子,包含翻译起始信号、信号肽、N端结构域以及该蛋白胶原样结构域的前七个胶原三聚体。四个短外显子(III至VI)编码该蛋白的胶原样结构域,外显子VII和VIII分别编码连接结构域和碳水化合物识别结构域。