miR-1343 attenuates pathways of fibrosis by targeting the TGF- receptors

Irreversible respiratory obstruction resulting from progressive airway damage, inflammation and fibrosis is a feature of several chronic respiratory diseases, including cystic fibrosis (CF), idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD). The cytokine transforming growth factor β (TGF-β) has a pivotal role in promoting lung fibrosis and is implicated in respiratory disease severity. In the present study, we show that a previously uncharacterized miRNA, miR-1343, reduces the expression of both TGF-β receptor 1 and 2 by directly targeting their 3′-UTRs. After TGF-β exposure, elevated intracellular miR-1343 significantly decreases levels of activated TGF-β effector molecules, pSMAD2 (phosphorylated SMAD2) and pSMAD3 (phosphorylated SMAD3), when compared with a non-targeting control miRNA. As a result, the abundance of fibrotic markers is reduced, cell migration into a scratch wound impaired and epithelial-to-mesenchymal transition (EMT) repressed. Mature miR-1343 is readily detected in human neutrophils and HL-60 cells and is activated in response to stress in A549 lung epithelial cells. miR-1343 may have direct therapeutic applications in fibrotic lung disease

Data from: The Caenorhabditis elegans Myc-Mondo/Mad complexes integrate diverse longevity signals

The Myc family of transcription factors regulates a variety of biological processes, including the cell cycle, growth, proliferation, metabolism, and apoptosis. In Caenorhabditis elegans, the “Myc interaction network” consists of two opposing heterodimeric complexes with antagonistic functions in transcriptional control: the Myc-Mondo:Mlx transcriptional activation complex and the Mad:Max transcriptional repression complex. In C. elegans, Mondo, Mlx, Mad, and Max are encoded by mml-1, mxl-2, mdl-1, and mxl-1, respectively. Here we show a similar antagonistic role for the C. elegans Myc-Mondo and Mad complexes in longevity control. Loss of mml-1 or mxl-2 shortens C. elegans lifespan. In contrast, loss of mdl-1 or mxl-1 increases longevity, dependent upon MML-1:MXL-2. The MML-1:MXL-2 and MDL-1:MXL-1 complexes function in both the insulin signaling and dietary restriction pathways. Furthermore, decreased insulin-like/IGF-1 signaling (ILS) or conditions of dietary restriction increase the accumulation of MML-1, consistent with the notion that the Myc family members function as sensors of metabolic status. Additionally, we find that Myc family members are regulated by distinct mechanisms, which would allow for integrated control of gene expression from diverse signals of metabolic status. We compared putative target genes based on ChIP-sequencing data in the modENCODE project and found significant overlap in genomic DNA binding between the major effectors of ILS (DAF-16/FoxO), DR (PHA-4/FoxA), and Myc family (MDL-1/Mad/Mxd) at common target genes, which suggests that diverse signals of metabolic status converge on overlapping transcriptional programs that influence aging. Consistent with this, there is over-enrichment at these common targets for genes that function in lifespan, stress response, and carbohydrate metabolism. Additionally, we find that Myc family members are also involved in stress response and the maintenance of protein homeostasis. Collectively, these findings indicate that Myc family members integrate diverse signals of metabolic status, to coordinate overlapping metabolic and cytoprotective transcriptional programs that determine the progression of aging

Raw Data of Mortality Observations of Replica Set Experiments

Raw Data of Mortality Observations of Replica Set Experiment

The <i>Caenorhabditis elegans</i> Myc-Mondo/Mad Complexes Integrate Diverse Longevity Signals

<div><p>The Myc family of transcription factors regulates a variety of biological processes, including the cell cycle, growth, proliferation, metabolism, and apoptosis. In <i>Caenorhabditis elegans,</i> the “Myc interaction network” consists of two opposing heterodimeric complexes with antagonistic functions in transcriptional control: the Myc-Mondo:Mlx transcriptional activation complex and the Mad:Max transcriptional repression complex. In <i>C. elegans,</i> Mondo, Mlx, Mad, and Max are encoded by <i>mml-1, mxl-2, mdl-1,</i> and <i>mxl-1,</i> respectively. Here we show a similar antagonistic role for the <i>C. elegans</i> Myc-Mondo and Mad complexes in longevity control. Loss of <i>mml-1</i> or <i>mxl-2</i> shortens <i>C. elegans</i> lifespan. In contrast, loss of <i>mdl-1</i> or <i>mxl-1</i> increases longevity, dependent upon MML-1:MXL-2. The MML-1:MXL-2 and MDL-1:MXL-1 complexes function in both the insulin signaling and dietary restriction pathways. Furthermore, decreased insulin-like/IGF-1 signaling (ILS) or conditions of dietary restriction increase the accumulation of MML-1, consistent with the notion that the Myc family members function as sensors of metabolic status. Additionally, we find that Myc family members are regulated by distinct mechanisms, which would allow for integrated control of gene expression from diverse signals of metabolic status. We compared putative target genes based on ChIP-sequencing data in the modENCODE project and found significant overlap in genomic DNA binding between the major effectors of ILS (DAF-16/FoxO), DR (PHA-4/FoxA), and Myc family (MDL-1/Mad/Mxd) at common target genes, which suggests that diverse signals of metabolic status converge on overlapping transcriptional programs that influence aging. Consistent with this, there is over-enrichment at these common targets for genes that function in lifespan, stress response, and carbohydrate metabolism. Additionally, we find that Myc family members are also involved in stress response and the maintenance of protein homeostasis. Collectively, these findings indicate that Myc family members integrate diverse signals of metabolic status, to coordinate overlapping metabolic and cytoprotective transcriptional programs that determine the progression of aging.</p></div

Loss of the Myc-Mondo complex impairs resistance to oxidative and thermal stress.

<p>(A) Loss of <i>mxl-2</i> impairs resistance to oxidative stress imposed by the exposure to <i>tert-</i>butylhydroperoxide (tBOOH) in wild-type and <i>daf-2(e1370)</i> mutant animals. Animals were exposed to 7.7mM tBOOH for the denoted time and survival was assessed. Loss of <i>mxl-2</i> in wild-type and <i>daf-2(e1370)</i> mutant animals impaired tBOOH survival to a similar extent. (B) Loss of <i>mxl-2</i> significantly impairs intrinsic thermotolerance (ITT) in wild-type and <i>daf-2(e1370)</i> mutant animals. (C) Loss of <i>mxl-2</i> does not further impair ITT in <i>daf-2(e1370);daf-16(mgDf47)</i> mutants. (D) <i>mxl-2</i> is dispensable for acquired thermotolerance in <i>daf-2(e1370)</i> mutants. Graphs were generated from the combined data from multiple experiments performed as described in Materials and Methods. Statistical analyses and experimental details can be found in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004278#pgen.1004278.s010" target="_blank">Dataset S3</a>.</p

Model for Myc-Mondo/Mad transcription factors in longevity control under basal conditions the Myc-Mondo activation complex (MML-1:MXL-2) is largely inactive, and transcription of genes encoding functions related to aging is limited by the Mad transcriptional repression complex (MDL-1:MXL-1).

<p>Conditions of reduced ILS and DR promote Myc-Mondo complex activity, likely by regulating cellular localization and transcription of <i>mml-1.</i> Myc-Mondo/Mad transcription factors may cooperate with DAF-16 and PHA-4 to modulate the expression of key metabolic and cytoprotective genes to influence aging.</p

The Myc-Mondo/Mad complexes intersect with dietary restriction in longevity.

<p>(A) RNAi inactivation of <i>pha-4</i> ablates the lifespan extension conferred by the <i>mxl-1(tm1530)</i> null mutation (light blue versus blue traces), and does not further shorten the lifespan of <i>mxl-2(tm1516)</i> null mutant animals (pink versus red traces). (B) RNAi inactivation of <i>mxl-2</i> partially suppresses the extended lifespan observed in <i>eat-2(ad465)</i> mutants (pink versus green traces). Similar results were obtained with <i>mml-1</i> RNAi (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004278#pgen.1004278.s008" target="_blank">Dataset S1</a>). (C) The <i>mxl-2(tm1516)</i> null mutation partially suppresses the extended lifespan observed in <i>eat-2(ad465)</i> mutant animals (pink versus green traces). (D) The <i>mxl-1(tm1530)</i> null mutation does not further extend the lifespan of <i>eat-2(ad465)</i> mutant animals (dark blue versus green traces). Similar results were obtained with <i>mxl-1</i> and <i>mdl-1</i> RNAi (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004278#pgen.1004278.s008" target="_blank">Dataset S1</a>). Traces represent the combined data from multiple separate trials. Complete information regarding the number of trials, total number of animals examined, and statistical significance for each experiment can be found in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004278#pgen.1004278.s008" target="_blank">Dataset S1</a>. Experiments shown within this figure were performed simultaneously with those shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004278#pgen-1004278-g002" target="_blank">Figure 2</a> and were split into multiple figures for readability.</p

ILS and DR pathways differentially regulate <i>mml-1</i> gene expression and MML-1 localization.

<p>(A) Nuclear accumulation of MML-1::GFP is increased in <i>daf-2(e1370)</i> and <i>eat-2(ad465)</i> mutant animals. Loss of <i>pha-4</i>, but not <i>daf-16</i>, blocks increases in MML-1::GFP nuclear accumulation in <i>daf-2(e1370)</i> and <i>eat-2(ad465)</i>, but does not affect MML-1::GFP localization under basal conditions. Representative images of Types I-V are found below the graph. Additional high-resolution images of MML-1::GFP expressing animals can be found in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004278#pgen.1004278.s011" target="_blank">File S1</a> (* denotes p < 0.05). (B) <i>mml-1</i> mRNA levels are significantly increased in <i>daf-2(e1370)</i>, but not in <i>mdl-1(tm311)</i> or <i>eat-2(ad465)</i> mutant animals, as compared to N2 (* denote p<0.05). (C) <i>mxl-2, mxl-1,</i> and <i>mdl-1</i> mRNA levels are unaffected in <i>daf-2(e1370)</i> mutants. A <i>daf-16</i> null mutation suppresses the increases in <i>mml-1</i> mRNA observed in <i>daf-2(e1370)</i> mutants. (* denotes p<0.01) (D) <i>mdl-1</i> mRNA levels are significantly increased in <i>daf-2(e1370);daf-16(mgDf47)</i> mutants (* denotes p<0.01).</p