An NCI-designated Comprehensive Cancer Center
By Krist Azizian | March 5, 2012

Krist AzizianDisrupting a critical molecular pathway for blood development reveals biology’s robustness.

Some faint at the sight of it. Others relish seeing it splattered gratuitously in vampire movies. Blood, with all its symbolism, has intrigued mankind for millennia. It is therefore unsurprising that blood’s development, called hematopoiesis, is well understood. Part of this understanding resulted from painstaking experimentation to study which genes control the potential for blood stem cells to mature, or differentiate, into the copious cell types that arise from relatively few precursors in the bone marrow. Differentiated cells then move on to function in vital processes such as carrying oxygen to tissues or fighting infections. Nevertheless, although the genes that govern hematopoiesis are mostly known, the exact molecular mechanisms that control how these genes work in concert remain elusive.

To shed light on these mechanisms, my colleagues and I utilized a pathway of gene regulation called RNA-interference (RNAi) to show that its disruption can influence the way that blood stem cells differentiate. One kind of RNAi important for hematopoiesis is the process by which short segments of RNA, called microRNAs (miRNAs), find and bind to messenger RNAs, longer segments that the cell uses to make proteins. This binding of miRNAs to their complementary messenger RNAs hinders protein synthesis and lowers the overall level of protein available to the cell for a given function. Fine-tuning of protein availability by miRNAs for when the cell needs it most is critical to maintaining health. When RNAi with miRNAs goes awry, protein levels become mismanaged and diseases such as cancer occur. Of interest then is whether or not blood stem cells strive to compensate for abnormal miRNA levels by modulating the amount of the enzymes that make them.

To study this, we used a type of RNA molecule called a short-hairpin to disrupt the production of the initial miRNA-processing enzyme, Drosha, in the nuclei of blood stem cells. We then tried to get these stem cells to differentiate into mature white blood cells called granulocytes and monocytes. This approach was powerful because disrupting Drosha meant that almost all miRNAs controlling hematopoiesis would be reduced, with those most important for differentiation likely persisting at relatively higher levels. It also meant that the stem cells, which rely heavily on miRNAs to direct other life-sustaining processes, should have died.

Surprisingly, quite the opposite happened. Not only did the cells lacking Drosha survive, but they also exhibited less apoptosis (the self-destruct program cells undergo when things get muddled). This was the first indication that some kind of compensation was occurring. Other observations suggesting compensation to combat Drosha’s deficiency, and the consequent lack of miRNA-controlled protein production, were an increase in the expression of downstream miRNA-processing enzymes and a shift of differentiation towards monocytes. Why monocytes seemed to be the mature cell of choice remains unclear, but activation of a default differentiation program to meet rising demand might be one reason. Finally, with Drosha sequestered, the persistence of one miRNA stood out above the rest, indicating its role as a master regulator of monocyte differentiation.

What this lends to our understanding of hematopoiesis is that mechanisms are ready during differentiation to help compensate for significant deficiencies in RNAi, thereby ensuring that blood development occurs. Because RNAi co-evolved with viruses as a form of defense against infection, it is likely that any compensation for its deficiencies resulted from evolution. If this is proven, it will add yet another level of intrigue to captivate hematologists for generations, or at least until the next blood-soaked horror flick.