With the new year comes a new round of government grants supporting the latest RNAi research. Among them is one that has been issued to advance the understanding of proteins that play an important role in the transmission of gene silencing information, and one that is funding experiments applying RNAi to the study of in vivo muscle metabolism.
The first grant was issued on Jan. 1 by the NIGMS, and is set to provide as much as $1.2 million over the next four years to Harvard University researcher Craig Hunter, who is investigating the subcellular localization, structure, activity, and regulation of the proteins SID-1 and SID-2 in C. elegans. According to the grant abstract, SID-1 has been identified as necessary for systemic RNAi. It is also predicted to contain 11 integral membrane domains and a large extracellular domain, both of which suggest that it may act as a channel or receptor for the uptake or transport of dsRNA.
Hunter recently co-authored a paper in Science stating that SID-1 is a multispan transmembrane protein that sensitizes Drosophila cells to soaking RNAi with a potency that was dependent on dsRNA length, and that SID-1 enables passive cellular uptake of dsRNA.
In the grant abstract, Hunter also noted that SID-2 has been identified as an integral membrane protein in C. elegans needed for the uptake of silencing information.
“To determine whether either SID protein acts as a channel or receptor, and to identify and characterize their presumed interactions with dsRNA, their transport activity will be investigated using nematode primary cell lines derived from wild-type and mutant embryos,” the abstract states. “To corroborate these findings, the transport activity of SID-1 and SID-2 expressed in heterologous systems will be assayed.”
Additionally, mouse SID-1 homologs will be expressed in nematodes and insect cell lines to determine if they can complement mutations of the sid-1 gene or perform any of the functions of SID-1. “Mouse embryonic stem cells will be characterized for expression of SID homologs and dsRNA uptake activity, as well as cell-to-cell spreading of silencing information,” the abstract states. “Finally, we will begin a functional analysis of the mouse homologs in ES cells and chimera embryos.”
“These studies may have a direct impact on the treatment of human genetic diseases and viral infection,” it concludes. “Although RNAi has shown to be effective, a major obstacle remains delivery of dsRNA into human cells both in culture and in vivo.”
The second grant, worth up to $336,000, was issued by the NIDDK to Joslin Diabetes Center researcher Laurie Goodyear. It runs from Jan 15 this year through November 2005.
This grant’s abstract explains that “skeletal muscle is the largest storage reservoir for glucose in the body, and defects in insulin action to promote muscle glucose uptake can result in elevations of circulating glucose concentrations and altered muscle glycogen concentrations.” However, the abstract notes, technical difficulties have hindered detailed research in this area.
The goals of Goodyear’s project are to develop a method for using RNAi in adult skeletal muscle in vivo, to determine if RNAi can be used to disrupt glucose transport and insulin action in this tissue, and to use RNAi to ameliorate skeletal muscle insulin resistance.
“Our studies demonstrate that electroporation is a reliable method for efficient transfection of skeletal muscle in vivo,” states the abstract. But to make the technique work, “we will need to make significant adjustments to the methodology.”
If successful, the project will “provide [a] ... valuable tool to study the molecular mechanisms regulating muscle metabolism and function in vivo, as well as for elucidating specific mechanisms underlying skeletal muscle insulin resistance,” the abstract concludes. “Ultimately, this approach could provide the basis for treatment of skeletal muscle insulin resistance and other chronic muscle diseases.”
—DM