The National Institutes of Health this month set aside more than $1 million to fund a handful of grants examining the roles of microRNAs in neurological dysfunction, immune responsiveness, and cancer, as well as the development of a new method for identifying targets of the non-coding RNAs.
The first grant was awarded to Shilpa Buch, a researcher at the University of Nebraska Medical Center, to support her efforts to uncover how miR-29b influences HIV-associated neuronal toxicity in opiate abusers.
In those infected with the virus, the use of opiates has been shown to cause increased neurologic and cognitive deficits, despite the availability of disease-controlling therapies, she noted her in grant's abstract.
In previous studies with morphine-dependent rhesus macaques with simian immunodeficiency virus, Buch and colleagues recapitulated the human syndrome demonstrating greater neuropathology and neuroinflammation, as well as rapid disease progression, compared with SIV-infected controls. “Mortality in these rapid progressors was associated with robust viral replication in both the periphery and the brain.”
With the NIH funding, Buch aims to use this established primate model to study how morphine impacts HIV disease outcome, specifically in terms of miRNA-mediated regulation of neurotoxicity. Her team has observed that in chronically SIV-infected macaques, morphine-mediated potentiation of neuropathogenesis correlated with dysregulation of miRNAs, with significant up-regulation of miR-29b — a miRNA that is also up-regulated in Alzheimer's disease and Parkinson's disease — in peripheral blood mononuclear cells and post-mortem brain tissue.
Buch will examine whether this neuropathogenesis involves miR-29b during the early stage of infection and whether the miRNA is critical for regulating genes controlling neuronal survival, such as the platelet-derived growth factor and its receptor, and results in cell death, according to the grant's abstract.
The project began on Feb. 1 and runs for five years. It is worth $436,367 in its first year.
Also receiving NIH funding is Emory University's Gary Bassell, who aims to further investigate his discovery of a novel molecular mechanism for neurotransmitter-regulated protein synthesis that involves “the release of a miRNA induced silencing complex from its target mRNA,” according to his grant's abstract.
Supported by the grant, Bassell and colleagues hope to determine whether this is a general mechanism by which miRNAs modulate activity-mediated mRNA translation in neurons, identify new miRNAs and targets that use the reversibility of miRISC to regulate activity-mediated translation, and uncover how different neurotransmitter signaling pathways might affect miRNA/RISC targeting to mRNAs.
Additionally, given the putative link between fragile X mental retardation and miRNAs, his team plans to assess whether phosphorylation of the fragile X mental retardation protein is capable of regulating a subset of miRNAs at synapses in a mouse model of the disease.
“This research has the potential to identify novel microRNAs regulated by neuronal activity, understand a unifying molecular mechanism for activity-regulated translation, and help to elucidate the pathophysiology of a neurological disorder at the molecular level,” he wrote in the grant's abstract.
The project, which is worth $234,000 in its first year, began on Feb. 1 and runs until Jan. 31, 2015.
The third grant went to Li-Fan Lu of the University of California, San Diego, to help finance his examination of miR-155, which has been linked to a variety of immune system processes, in repressing SOCS1, a negative regulator of cytokine signaling that is required for regulatory T cell, or Treg, function.
In previous studies, Lu found that Foxp3, a transcription factor key to controlling Tregs, drives high expression of miR-155 and “promotes the competitive fitness of Treg cells by inducing SOCS1 down-regulation,” according to his grant's abstract. “Subsequent studies conducted by other groups have implied a role for the miR-155/SOCS1 axis in tuning macrophage responsiveness to [lipopolysaccharide]-induced endotoxin tolerance, as well as cytokine production by dendritic cells.”
But because miRNAs target a range of mRNAs, immune responses identified in miR-155-deficient mice may be attributed to genes other than SOCS1, Lu noted.
To address this issue, he and his team intend to use the NIH funding to identify miR-155's full role in SOCS1 regulation.
“First, we will examine the combined effects of SOCS1 deficiency/haploinsufficiency and miR-155 deficiency to see if aspects of the observed miR-155 phenotype are reversed by loss of SOCS1 function,” he wrote in the grant's abstract. “Next, by generating a new mouse model with mutations specifically disrupting the interaction between miR-155 and SOCS1 gene and by comparing these mice to miR-155-deficient mice, we will be able to isolate the effects of miR-155 on a single target and to explore the biological significance of SOCS1 repression in miR-155-mediated immune regulation.”
He added that, because miR-155 has been implicated in the inflammatory responses required for autoimmunity, the investigators plan to examine SOCS1 repression mediated by the miRNA in an animal model of multiple sclerosis.
The grant project began on Feb. 1 and runs for three years. It is worth $193,750 in the first year.
At Northwestern University, researcher Jian-Jun Wei also received an NIH grant to support his investigation into miR-182 and high-grade serous carcinoma, an aggressive form of ovarian cancer.
Mutations in BRCA1 and BRCA2 are a key risk factor in serous carcinoma, and the discovery that miR-182 represses BRCA1 expression suggests that the miRNA, which is over-expressed in cells with DNA damage, may play a part in the disease, according to the grant's abstract.
In light of this, Wei and his colleagues hypothesize that miR-182 over-expression is an “early genetic event and an important trigger of tumorigenesis” in the cancer through dysregulation of associated genes.
To test this, they will use the NIH funding to examine how miR-182 responds to DNA damage in fallopian tube secretory epithelial cells, in which ovarian cancer can originate, and whether IR-induced miR-182 over-expression requires a mutant form of the tumor suppressor p53.
The team will also examine the kinetics of DNA damage repair in these cells in the presence and absence of miR-182, and look into whether miR-182-mediated tumorigenesis can be enhanced by negative regulation of two of its predicted target genes, according to the grant's abstract.
The investigators then plan to see if miR-182 over-expression in the fallopian tube cells triggers tumor growth in vitro and if its knockdown can reduce tumor burden and metastasis in vivo.
The grant, which began on Feb. 1 and runs for three years, is worth $151,217 in the first year.
The last grant was awarded to Oregon Health & Science University investigator Richard Goodman, who is refining a miRNA target-identification method that uses an epitope-tagged, dominant negative version of the RISC component GW182 to purify miRNA/mRNA complexes prior to miRNA-mediated mRNA degradation.
The method, dubbed RISC-Trap, was described late last year in the Proceedings of the National Academy of Sciences.
According to the grant's abstract, the method enables researchers to “address fundamental problems in microRNA biology, such as, what proportion of mRNA targets undergo degradation versus translational arrest, which targets are direct or indirect, and whether some interactions occur specifically in neural versus non-neural cells.”
With the NIH funding, Goodman and his team will use RISC-Trap to determine in vitro which targets of miR-124 targets are regulated by mRNA degradation versus translational arrest. “These studies will provide a comprehensive picture of the regulatory effects of an important brain microRNA and a straightforward and easily applicable method for identifying microRNA targets in general,” he wrote.
Next, the investigators plan to identify miR-124 targets specific to neuronal cells and determine if their mode of action differs from targets not specific to such cells, with the goal of demonstrating RISC-Trap's usefulness in uncovering new aspects of miRNA function.
The grant project began on Feb. 15 and runs until Jan. 31, 2015. It is worth $231,000 in its first year.