ABSTRACT
The adaptive cellular response to low oxygen tensions is mediated by the hypoxia-inducible factors (HIFs), a family of heterodimeric transcription factors composed of HIF-α and HIF-ß subunits. Prolonged HIF expression is a key contributor to cellular transformation, tumorigenesis and metastasis. As such, HIF degradation under hypoxic conditions is an essential homeostatic and tumour-suppressive mechanism. LIMD1 complexes with PHD2 and VHL in physiological oxygen levels (normoxia) to facilitate proteasomal degradation of the HIF-α subunit. Here, we identify LIMD1 as a HIF-1 target gene, which mediates a previously uncharacterised, negative regulatory feedback mechanism for hypoxic HIF-α degradation by modulating PHD2-LIMD1-VHL complex formation. Hypoxic induction of LIMD1 expression results in increased HIF-α protein degradation, inhibiting HIF-1 target gene expression, tumour growth and vascularisation. Furthermore, we report that copy number variation at the LIMD1 locus occurs in 47.1% of lung adenocarcinoma patients, correlates with enhanced expression of a HIF target gene signature and is a negative prognostic indicator. Taken together, our data open a new field of research into the aetiology, diagnosis and prognosis of LIMD1-negative lung cancers.
Subject(s)
Adenocarcinoma/genetics , Gene Expression Regulation, Neoplastic , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Intracellular Signaling Peptides and Proteins/metabolism , LIM Domain Proteins/metabolism , Lung Neoplasms/genetics , Adenocarcinoma/diagnosis , Adenocarcinoma/metabolism , Adenocarcinoma/mortality , Adult , Aged , Aged, 80 and over , Animals , Carcinogenesis/genetics , Carcinogenesis/metabolism , Cell Hypoxia/genetics , Cell Hypoxia/physiology , Cell Line, Tumor , Feedback, Physiological , Female , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Intracellular Signaling Peptides and Proteins/genetics , LIM Domain Proteins/genetics , Lung Neoplasms/diagnosis , Lung Neoplasms/metabolism , Lung Neoplasms/mortality , Male , Mice , Middle Aged , Prognosis , Survival Analysis , Vascular Endothelial Growth Factor A/geneticsABSTRACT
As core components of the microRNA-induced silencing complex (miRISC), Argonaute (AGO) proteins interact with TNRC6 proteins, recruiting other effectors of translational repression/mRNA destabilization. Here, we show that LIMD1 coordinates the assembly of an AGO-TNRC6 containing miRISC complex by binding both proteins simultaneously at distinct interfaces. Phosphorylation of AGO2 at Ser 387 by Akt3 induces LIMD1 binding, which in turn enables AGO2 to interact with TNRC6A and downstream effector DDX6. Conservation of this serine in AGO1 and 4 indicates this mechanism may be a fundamental requirement for AGO function and miRISC assembly. Upon CRISPR-Cas9-mediated knockout of LIMD1, AGO2 miRNA-silencing function is lost and miRNA silencing becomes dependent on a complex formed by AGO3 and the LIMD1 family member WTIP. The switch to AGO3 utilization occurs due to the presence of a glutamic acid residue (E390) on the interaction interface, which allows AGO3 to bind to LIMD1, AJUBA, and WTIP irrespective of Akt signaling.
