Novel MicroRNA Candidates and miRNA-mRNA Pairs in Embryonic Stem (ES) Cells

MicroRNAS (miRNAS: a class of short non-coding RNAs) are emerging as important agents of post transcriptional gene regulation and integral components of gene networks. MiRNAs have been strongly linked to stem cells, which have a remarkable dual role in development. They can either continuously replenish themselves (self-renewal), or differentiate into cells that execute a limited number of specific actions (pluripotence).In order to identify novel miRNAs from narrow windows of development we carried out an in silico search for micro-conserved elements (MCE) in adult tissue progenitor transcript sequences. A plethora of previously unknown miRNA candidates were revealed including 545 small RNAs that are enriched in embryonic stem (ES) cells over adult cells. Approximately 20% of these novel candidates are down-regulated in ES (Dicer(-/-)) ES cells that are impaired in miRNA maturation. The ES-enriched miRNA candidates exhibit distinct and opposite expression trends from mmu-mirs (an abundant class in adult tissues) during retinoic acid (RA)-induced ES cell differentiation. Significant perturbation of trends is found in both miRNAs and novel candidates in ES (GCNF(-/-)) cells, which display loss of repression of pluripotence genes upon differentiation.Combining expression profile information with miRNA target prediction, we identified miRNA-mRNA pairs that correlate with ES cell pluripotence and differentiation. Perturbation of these pairs in the ES (GCNF(-/-)) mutant suggests a role for miRNAs in the core regulatory networks underlying ES cell self-renewal, pluripotence and differentiation

LFA-1 Affinity Regulation Is Necessary for the Activation and Proliferation of Naive T Cells*S⃞

The activation of LFA-1 (lymphocyte function-associated antigen) is a critical event for T cell co-stimulation. The mechanism of LFA-1 activation involves both affinity and avidity regulation, but the role of each in T cell activation remains unclear. We have identified antibodies that recognize and block different affinity states of the mouse LFA-1 I-domain. Monoclonal antibody 2D7 preferentially binds to the low affinity conformation, and this specific binding is abolished when LFA-1 is locked in the high affinity conformation. In contrast, M17/4 can bind both the locked high and low affinity forms of LFA-1. Although both 2D7 and M17/4 are blocking antibodies, 2D7 is significantly less potent than M17/4 in blocking LFA-1-mediated adhesion; thus, blocking high affinity LFA-1 is critical for preventing LFA-1-mediated adhesion. Using these reagents, we investigated whether LFA-1 affinity regulation affects T cell activation. We found that blocking high affinity LFA-1 prevents interleukin-2 production and T cell proliferation, demonstrated by TCR cross-linking and antigen-specific stimulation. Furthermore, there is a differential requirement of high affinity LFA-1 in the activation of CD4+ and CD8+ T cells. Although CD4+ T cell activation depends on both high and low affinity LFA-1, only high affinity LFA-1 provides co-stimulation for CD8+ T cell activation. Together, our data demonstrated that the I-domain of LFA-1 changes to the high affinity state in primary T cells, and high affinity LFA-1 is critical for facilitating T cell activation. This implicates LFA-1 activation as a novel regulatory mechanism for the modulation of T cell activation and proliferation

Field assessment of the risk of feral cat baits to non-target species in eastern Australia

ABSTRACT Feral cats pose a significant threat to wildlife, agriculture and human health through predation, disease transmission and competition with native animals. Controlling feral cats and their impacts, however, is challenging. New and emerging 1080-based feral cat baits have shown promising results in western and central Australia, however the safety of these new baits for non-target species in eastern Australia, where many native animals are more sensitive to 1080 than their western conspecifics, has not been assessed. We investigated the uptake by non-target animals of 499 toxic Eradicat® baits across five different eastern Australian environs, and the uptake of non-toxic Eradicat® and Hisstory® baits at an additional two sites. Using field-based observations of species eating or removing baits, we determined that 13 non-target species (eight mammals, four birds, one reptile) were at high risk of individual mortality, with individuals of 11 of those 13 species (four birds, seven mammals) observed consuming enough toxic Eradicat® in a single visit to ingest a lethal dose of 1080. Feral cats (the target species) consumed only 3.1% of monitored baits, which was only 52% of the 31 baits they encountered. We recommend undertaking targeted population monitoring of species identified at high risk of individual mortality, to determine whether Eradicat® baits present a population-level risk to these species. Our findings suggest that the small-sized Eradicat® baits present a greater risk to non-target species in eastern Australia than the larger traditional 1080-based meat baits used for the control of wild dogs and foxes. Our study highlights the importance of performing risk assessments for different bait types, even when the same toxin is used, and of performing site-specific non-target risk assessments of new baits such as Eradicat® to assist developing guidelines for their safe and effective use in different environs. This article is protected by copyright. All rights reserved