Genome-wide screening of NEAT1 regulators reveals cross-regulation between paraspeckles and mitochondria

Abstract

The long noncoding RNA NEAT1 (nuclear enriched abundant transcript 1) nucleates the formation of paraspeckles, which constitute a type of nuclear body with multiple roles in gene expression. Here we identify NEAT1 regulators using an endogenous NEAT1 promoter-driven enhanced green fluorescent protein reporter in human cells coupled with genome-wide RNAi screens. The screens unexpectedly yield gene candidates involved in mitochondrial functions as essential regulators of NEAT1 expression and paraspeckle formation. Depletion of mitochondrial proteins and treatment of mitochondrial stressors both lead to aberrant NEAT1 expression via ATF2 as well as altered morphology and numbers of paraspeckles. These changes result in enhanced retention of mRNAs of nuclear-encoded mitochondrial proteins (mito-mRNAs) in paraspeckles. Correspondingly, NEAT1 depletion has profound effects on mitochondrial dynamics and function by altering the sequestration of mito-mRNAs in paraspeckles. Overall, our data provide a rich resource for understanding NEAT1 and paraspeckle regulation, and reveal a cross-regulation between paraspeckles and mitochondria.

Forced isoform switching of Neat1_1 to Neat1_2 leads to the loss of Neat1_1 and the hyperformation of paraspeckles but does not affect the development and growth of mice

“…This hypothesis is consistent with a recent report showing that deletion of PAS in (Nakagawa et al 2011;Nakagawa et al 2014). Indeed, the expression of Neat1_2 and paraspeckle formation are enhanced by various stimuli, including viral infection (Saha et al 2006), prolonged cell culture (Nakagawa et al 2011), proteasome inhibition ), poly-I:C transfection (Imamura et al 2014), and mitochondrial stress (Wang et al 2018). Importantly, Neat1 is controlled by the p53 pathway, but cancer progression is enhanced or inhibited in a context-dependent manner: the lack of Neat1 enhanced the development of premalignant pancreatic intraepithelial neoplasias in KrasG12D-expressing mice (Mello et al 2017), whereas the progression of skin neoplasias induced by carcinogens was suppressed in Neat1 KO mice (Adriaens et al 2016).…”

“…In this study, we forced the isoform switching of Neat1 by deleting the PAS required for the production of Neat1_1, which has been shown to increase the expression of Neat1_2 at the expense of Neat1_1 expression (Li et al 2017;Wang et al 2018;Yamazaki et al 2018;Modic et al 2019). Despite the lack of Neat1_1 expression and hyperformation of paraspeckles in various tissues and cells, no overt phenotype was observed in the mutant mice lacking the Neat1 PAS (Neat1 Δ PAS/ Δ PAS ).…”

NEAT1 is overexpressed in Parkinson's disease substantia nigra and confers drug‐inducible neuroprotection from oxidative stress

“…3 E ). Because NEAT1 has recently been implicated in the regulation of mitochondrial formation and function (23), we also quantified mitochondria under conditions of NEAT1 knockdown in HEK-293T cells. Microscopic quantification of the mitochondrial-selective fluorescent probe MitoTracker (40) and comparative qPCR using 2 mitochondrial DNA–specific primers demonstrated decreased mitochondrial abundance (Supplemental Fig.…”

“…The proposed regulation by mitochondria over NEAT1 expression and paraspeckle formation is complex and multilayered; although mitochondrial stressors change the levels of both variants, knockdown of specific mitochondrial genes affects the intervariant balance as well (23). An interaction between mitochondria abundance and function and the bioavailability of mitochondrial proteins may thus explain the unique up-regulation of NEAT1 and nuclear paraspeckles in PD and Huntington's disease.…”

“…Although the sequence of NEAT1 is not highly conserved across species, its functions as the paraspeckle assembly scaffold are well established (22). Notably, NEAT1 also affects mitochondrial abundance and function (23). Nuclear paraspeckles have been identified as causally involved in immune responses (24) and cancer (25), and recent research on NEAT1 functions in the context of the CNS has revealed its involvement in Huntington’s disease (26) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)–induced neurotoxicity (27, 28).…”