MicroRNAs and hematopoietic differentiation

描述（由申请人提供）：理解和操纵造血分化需要了解在这一过程中协调基因表达程序的调控电路。一类基因调控分子是microRNAs （miRNAs）——一种微小的内源性rna，长度约为22个核苷酸，被认为利用rna干扰途径的元件转录后下调蛋白质编码基因的表达。从mirna在造血分化过程中发挥重要调节作用的假设开始，Lodish和Bartel实验室合作从小鼠骨髓中克隆了大约100种不同的mirna。这些包括五种被称为“造血mirna”的mirna，因为它们在造血细胞系中高度或优先表达。这5个基因中有3个也来自于先前与白血病相关的染色体断点或畸变相关的基因座。初步研究表明，在骨髓祖细胞中，其中一种mirna的异位表达调节了细胞培养和移植小鼠的造血分化。

DESCRIPTION (provided by applicant): Understanding and manipulating hematopoietic differentiation requires knowing the regulatory circuitry that orchestrates the programs of gene expression during this process. One class of gene regulatory molecules are the microRNAs (miRNAs) - tiny endogenous RNAs, about 22 nucleotides in length, that are thought to use the elements of the RNA-interference pathway to post transcriptionally down-regulate the expression of protein-coding genes. Starting with the hypothesis that miRNAs are playing important regulatory roles during hematopoietic differentiation, the Lodish and Bartel labs have collaborated to clone about 100 different miRNAs from mouse bone marrow. These include five miRNAs referred to as "hematopoietic miRNAs", because they are highly or preferentially expressed in hematopoietic cell lineages. Three of the five also derive from loci associated with chromosomal breakpoints or aberrations previously linked to leukemias. Preliminary studies show that ectopic expression of one of these miRNAs in bone marrow progenitors modulates hematopoietic differentiation both in cell culture and in transplanted mice. The experiments of this proposal focus on the hematopoietic miRNAs with the broad, long-term objective of understanding the gene regulatory events needed for hematopoietic stem cell and progenitor maintenance and differentiation. The specific aims are: 1) To examine the consequences of altered miRNA expression during hematopoiesis. 2) To identify the regulatory targets of hematopoietic miRNAs and examine the consequences of disrupting miRNA regulation of these targets. 3) To identify additional hematopoietic miRNAs. These experiments include the ectopic expression of hematopoietic miRNAs in hematopoietic stem cells and lineage-committed progenitors, knock-outs of miRNA genes, in vitro validation of predicted miRNA regulatory targets, in vivo substitution of target genes with versions unresponsive to miRNA regulation, cloning of additional miRNAs from hematopoietic tissues, and further expression analyses. They seek to place miRNAs within specific gene regulatory pathways needed for hematopoietic stem cell maintenance and lymphoid and myeloid differentiation. They will also address more fundamental issues regarding miRNA regulation, such as the combinatorial control of expression by miRNAs and the features of functional miRNA complementary sites within vertebrate mRNAs. Thus, these experiments will provide important insights for understanding the action of miRNAs in mammals and how their dysfunction might contribute to both hematological and other human diseases.