In this experimental setting, calcineurin blockade with CsA prevented LV hypertrophy due to pressure overload. TAB mice treated with CsA maintain normal LV size and systolic function.

We describe the isolation and characterization of ICL1 from the rice blast fungus Magnaporthe grisea, a gene that encodes isocitrate lyase, one of the principal enzymes of the glyoxylate cycle. ICL1 shows elevated expression during development of infection structures and cuticle penetration, and a targeted gene replacement showed that the gene is required for full virulence by M. grisea. In particular, we found that the prepenetration stage of development, before entry into plant tissue, is affected by loss of the glyoxylate cycle. There is a delay in germination, infection-related development and cuticle penetration in Delta icl1 mutants. Recent reports have shown the importance of the glyoxylate cycle in the virulence of the human pathogenic fungus Candida albicans and the bacterial pathogen Mycobacterium tuberculosis. Our results indicate that the glyoxylate cycle is also important in this plant pathogenic fungus, demonstrating the widespread utility of the pathway in microbial pathogenesis.

The Magnaporthe oryzae avirulence gene AvrPiz-t activates immunity in a gene-for-gene fashion to rice mediated by the blast resistance gene Piz-t. To dissect the molecular mechanism underlying their recognition, we initiated the cloning of AvrPiz-t using a map-based cloning strategy. The AvrPiz-t gene was delimited to an approximately 21-kb genomic fragment, in which six genes were predicted. Complementation tests of each of these six candidate genes led to the final identification of AvrPiz-t, which encodes a 108-amino-acid predicted secreted protein with unknown function and no homologues in M. oryzae or in other sequenced fungi. We found that AvrPiz-t is present in the virulent isolate GUY11 but contains a Pot3 insertion at a position 462 bp upstream from the start codon. Complementation tests of AvrPiz-t genes driven by promoters of varying length revealed that a promoter larger than 462 bp is essential to maintain the AvrPiz-t function. These results suggest that a Pot3 insertion in GUY11 might interfere with the proper function of AvrPiz-t. Additionally, we found that AvrPiz-t can suppress the programmed cell death triggered by mouse BAX protein in Nicotiana benthamiana, identifying a mechanism by which AvrPiz-t may contribute virulence of M. oryzae.

A previous study identified MoRgs1 as an RGS protein that negative regulates G-protein signaling to control developmental processes such as conidiation and appressorium formation in Magnaporthe oryzae. Here, we characterized additional seven RGS and RGS-like proteins (MoRgs2 through MoRgs8). We found that MoRgs1 and MoRgs4 positively regulate surface hydrophobicity, conidiation, and mating. Indifference to MoRgs1, MoRgs4 has a role in regulating laccase and peroxidase activities. MoRgs1, MoRgs2, MoRgs3, MoRgs4, MoRgs6, and MoRgs7 are important for germ tube growth and appressorium formation. Interestingly, MoRgs7 and MoRgs8 exhibit a unique domain structure in which the RGS domain is linked to a seven-transmembrane motif, a hallmark of G-protein coupled receptors (GPCRs). We have also shown that MoRgs1 regulates mating through negative regulation of Gα MoMagB and is involved in the maintenance of cell wall integrity. While all proteins appear to be involved in the control of intracellular cAMP levels, only MoRgs1, MoRgs3, MoRgs4, and MoRgs7 are required for full virulence. Taking together, in addition to MoRgs1 functions as a prominent RGS protein in M. oryzae, MoRgs4 and other RGS and RGS-like proteins are also involved in a complex process governing asexual/sexual development, appressorium formation, and pathogenicity.

L-type Ca2+ channels are characterized by their unique sensitivity to organic Ca2+ channel modulators like the 1,4-dihydropyridines (DHPs). To identify molecular motifs mediating DHP sensitivity, we transferred this sensitivity from L-type Ca2+ channels to the DHP-insensitive class A brain Ca2+ channel, BI-2. Expression of chimeras revealed minimum sequence stretches conferring DHP sensitivity including segments IIIS5, IIIS6, and the connecting linker, as well as the IVS5-IVS6 linker plus segment IVS6. DHP agonist and antagonist effects are determined by different regions within the repeat IV motif. Sequence regions responsible for DHP sensitivity comprise only 9.4% of the overall primary structure of a DHP-sensitive alpha 1A/alpha 1S construct. This chimera fully exhibits the DHP sensitivity of channels formed by L-type alpha 1 subunits. In addition, it displays the electrophysiological properties of alpha 1A, as well as its sensitivity toward the peptide toxins omega-agatoxin IVA and omega-conotoxin MVIIC.

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Neural progenitor cell (NPC) therapy is considered a promising treatment modality for multiple sclerosis (MS), potentially acting through neural repair. Here, we showed that intravenous administration of NPCs ameliorated experimental autoimmune encephalomyelitis (EAE) by selectively inhibiting pathogenic T helper 17 (Th17) cell differentiation. Leukemia inhibitory factor (LIF) produced by NPCs was responsible for the observed EAE suppression. Through the inducible LIF receptor expression, LIF inhibited the differentiation of Th17 cells in EAE mice and that from MS subjects. At the molecular level, LIF exerted an opposing effect on interleukin 6 (IL-6)-induced signal transducer and activator of transcription 3 (STAT3) phosphorylation required for Th17 cell differentiation by triggering a signaling cascade that activated extracellular signal-regulated MAP kinase (ERK) and upregulated suppressor of cytokine signaling 3 (SOCS3) expression. This study reveals a critical role for LIF in regulating Th17 cell differentiation and provides insights into the mechanisms of action of NPC therapy in MS.

SummaryMagnaporthe grisea, the causal agent of rice blast disease, invades plant tissue due to the action of specialized infection structures called appressoria, which are used to breach the leaf cuticle and allow development of intracellular, infectious hyphae. In this report we demonstrate that peroxisomal carnitine acetyl transferase (CAT) activity is necessary for appressorium function, and in particular, for the elaboration of primary penetration hyphae. The major CAT activity in M. grisea is encoded by the PTH2 gene, which shows elevated expression in response to acetate and lipid, and is regulated by the cyclic AMP response pathway. Furthermore, a Pth2-GFP fusion protein colocalizes with a peroxisomal marker protein. Targeted deletion of PTH2, generated mutants that were completely non-pathogenic, lacked CAT activity and were unable to utilize a range of lipid substrates. The impairment of appressorium function in Dpth2 was associated with a delay in lipid reserve mobilization from germ tubes into developing infection cells, and abnormal chitin distribution in infection structures. Addition of glucose to Dpth2 mutants partially restored the ability to cause rice blast disease and lipid reserve mobilization. Taken together, our findings provide evidence that Pth2 plays a role in the generation of acetyl CoA pools necessary for appressorium function and rapid elaboration of penetration hyphae during host infection.