MiR-35 buffers apoptosis thresholds in the C. elegans germline by antagonizing both MAPK and core apoptosis pathways

Anh T Tran,Samuel J Krempel,Eric M Chapman,Matthew Eroglu,Thomas F Duchaine,Bin Yu,Mathieu N Flamand,W Brent Derry

doi:10.1038/s41418-019-0325-6

Abstract

Apoptosis is a genetically programmed cell death process with profound roles in development and disease. MicroRNAs modulate the expression of many proteins and are often deregulated in human diseases, such as cancer. C. elegans germ cells undergo apoptosis in response to genotoxic stress by the combined activities of the core apoptosis and MAPK pathways, but how their signalling thresholds are buffered is an open question. Here we show mir-35–42 miRNA family play a dual role in antagonizing both NDK-1, a positive regulator of MAPK signalling, and the BH3-only pro-apoptotic protein EGL-1 to regulate the magnitude of DNA damage-induced apoptosis in the C. elegans germline. We show that while miR-35 represses EGL-1 by promoting transcript degradation, repression of NDK-1 may be through sequestration of the transcript to inhibit translation. Importantly, dramatic increase in NDK-1 expression was observed in cells about to die. In the absence of miR-35, increased NDK-1 activity enhanced MAPK signalling that lead to significant increases in germ cell death. Our findings demonstrate that NDK-1 acts upstream of (or in parallel to) EGL-1, and that miR-35 targets both egl-1 and ndk-1 to fine-tune cell killing in response to genotoxic stress.

Highlights

Ablation of ndk-1 resulted in no difference in p-MPK-1 compared with controls, but a twofold reduction in p-MPK-1 was observed in mir-35 mutants after ablation of ndk-1
We found that elevated MPK-1 phosphorylation in the pachytene region of mir-35 mutants was the result of increased NDK-1 activity, indicating it can stimulate MPK-1-dependent apoptosis in the germline
Since increased MPK-1 phosphorylation was evident in both the absence and presence of IR, but there was no increase in physiological apoptosis, we suggest that the miR-35 family acts to buffer mitogen-activated protein kinase (MAPK) activity under normal and stressed conditions

Summary

Present address

Absence of mir-35-41 results in up to fourfold more cell killing in response to DNA damage compared with wildtype (N2) animals This is accomplished by coordinately inhibiting both NDK-1, a positive regulator of MAPK signalling, and the BH3-only pro-apoptotic protein EGL-1. To determine the effect of genotoxic stress on the C. elegans germline in the absence of miR-35 activity, we exposed mir-35(gk262) mutants to increasing doses of ionizing irradiation (IR) and quantified germ cell corpses. Ablation of ndk-1 resulted in no difference in p-MPK-1 compared with controls, but a twofold reduction in p-MPK-1 was observed in mir-35 mutants after ablation of ndk-1

60 Gy egl-1

Discussion

Materials and methods

Findings

Compliance with ethical standards