Cloning and tissue-specific expression of mouse macrophage colony-stimulating factor mRNA.
Cloning and tissue-specific expression of mouse macrophage colony-stimulating factor mRNA.
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小鼠巨噬细胞集落刺激因子 mRNA 的克隆和组织特异性表达。
DOI：
10.1073/pnas.84.5.1157
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发表时间：
1987
影响因子：
11.1
通讯作者：
Lusis,AJ
中科院分区：
文献类型：
--
作者：
Rajavashisth,TB;Eng,R;Shadduck,RK;Waheed,A;Ben-Avram,CM;Shively,JE;Lusis,AJ
巨噬细胞集落刺激因子（CSF-1）刺激骨髓祖细胞产生巨噬细胞。我们通过抗体筛选表达载体λ gt 11中的小鼠L细胞cDNA文库，鉴定了鼠CSF-1的cDNA克隆。约150，000个重组噬斑的筛选产生6个克隆，其与针对变性和还原的小鼠L细胞CSF-1产生的抗体反应良好。基于CSF-1的氨基末端氨基酸序列，用合成的寡核苷酸进一步筛选这些克隆。对与寡核苷酸杂交的一个克隆进行测序，发现其含有单个开放阅读框。这包括成熟蛋白的68个氨基酸，包括我们以前报道的整个氨基末端序列。在此之前，似乎是一个31个氨基酸的信号肽。印迹分析表明，该cDNA杂交到约4.5 kb的主要mRNA种类以及小鼠L细胞中存在的几个较小的、丰度较低的mRNA种类（3.8、2.3和1.4 kb）。用来自产生CSF-1的人胰腺癌细胞系的mRNA观察到类似的杂交模式。在各种小鼠组织中观察到CSF-1的mRNA种类的定性和定量表达的显著差异。肝脏主要表达1.4-kb的物质，心脏和肺主要表达4.5-kb的物质，大脑表达高水平的4.5-kb和1.4-kb的物质，肠道缺乏可检测的CSF-1转录物。Southern印迹分析表明，CSF-1基因是目前作为一个单一的副本在小鼠单倍体基因组中，它不是重排或扩增L细胞。
Macrophage colony-stimulating factor (CSF-1) stimulates the production of macrophages from bone marrow progenitor cells. We have identified a cDNA clone for murine CSF-1 by antibody screening of a mouse L-cell cDNA library in the expression vector lambda gt11. A screen of about 150,000 recombinant plaques yielded 6 clones that reacted well with an antibody raised against denatured and reduced mouse L-cell CSF-1. These clones were further screened with synthetic oligonucleotides based on the amino-terminal amino acid sequence of CSF-1. One clone, which hybridized to the oligonucleotides, was sequenced and found to contain a single open reading frame. This encompassed 68 amino acids of the mature protein, including the entire amino-terminal sequence we previously reported. This is preceded by what appears to be a 31 amino acid signal peptide. Blot analysis showed that this cDNA hybridizes to a major mRNA species of about 4.5 kilobases (kb) as well as several smaller, less abundant mRNA species (3.8, 2.3, and 1.4 kb) present in mouse L cells. A similar pattern of hybridization was observed with mRNA from a human pancreatic carcinoma cell line that produces CSF-1. Striking differences in the qualitative and quantitative expression of mRNA species for CSF-1 were observed in various mouse tissues. Liver expressed primarily a 1.4-kb species, heart and lung expressed primarily a 4.5-kb species, brain expressed high levels of both the 4.5-kb and 1.4-kb species, and intestine lacked detectable CSF-1 transcripts. Southern blot analysis suggests that the CSF-1 gene is present as a single copy in the mouse haploid genome and that it is not rearranged or amplified in L cells.