Divergent roles of ALS-linked proteins FUS/TLS and TDP-43 intersect in processing long pre-mRNAs

Abstract

FUS/TLS (fused in sarcoma/translocated in liposarcoma) and TDP-43 are integrally involved in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. We found that FUS/TLS binds to RNAs from >5,500 genes in mouse and human brain, primarily through a GUGGU-binding motif. We identified a sawtooth-like binding pattern, consistent with co-transcriptional deposition of FUS/TLS. Depletion of FUS/TLS from the adult nervous system altered the levels or splicing of >950 mRNAs, most of which are distinct from RNAs dependent on TDP-43. Abundance of only 45 RNAs was reduced after depletion of either TDP-43 or FUS/TLS from mouse brain, but among these were mRNAs that were transcribed from genes with exceptionally long introns and that encode proteins that are essential for neuronal integrity. Expression levels of a subset of these were lowered after TDP-43 or FUS/TLS depletion in stem cell-derived human neurons and in TDP-43 aggregate–containing motor neurons in sporadic ALS, supporting a common loss-of-function pathway as one component underlying motor neuron death from misregulation of TDP-43 or FUS/TLS.

Linking RNA Dysfunction and Neurodegeneration in Amyotrophic Lateral Sclerosis

“…A minor portion of the genes most prominently downregulated by TDP43 depletion were affected in a similar fashion by FUS knockdown, including neuroligins and neurexins that are essential for neuronal health. Still, genes displaying overlapping patterns of dysregulation represented only a small fraction of the total, with the majority of differentially expressed genes being distinct in each case [36]. In addition, targeted reduction in TDP43 or FUS produced disparate effects in NSC34 mouse motor neuron-like cells [62].…”

“…TDP43 binds to and regulates thousands of RNA targets [13,14,[36][37][38], and is thus in a unique position to dramatically affect gene expression on a global scale. The RNA-binding domains of TDP43 and FUS are essential for toxicity in ALS model systems, testifying to the importance of RNA dysregulation in ALS pathogenesis [34,[39][40][41].…”

“…Most ALS-associated mutations in FUS involve the nuclear localization signal [15,16], resulting in deficient nuclear import and cytoplasmic accumulation of the protein [61]. Reductions in FUS induce widespread changes in gene expression and splicing that demonstrate partial overlap with those observed in TDP43-deficient murine primary neurons and human induced pluripotent stem cell (iPSC)-derived neurons [36,59]. Although TDP43 and FUS display distinct RNA binding patterns, they both strongly affect the expression of genes with long (>150 Kbp) introns.…”

“…TDP43 exhibits a conserved RNA binding pattern in human neuroblastoma cell lines (SH-SY5Y), human embryonic stem cells, and postmortem human cortex [13]. Likewise, the RNA targets of FUS are 70 % conserved in mouse and human brain [36]. Despite the distinct cellular heterogeneity typical of both motor cortex and iPSC cultures, several differentially expressed genes were commonly dysregulated in human iPSC-derived motor neurons and motor cortex from ALS patients with C9orf72 expansion mutations [68].…”

“…1). ASO-mediated knockdown of TDP43 altered 779 splicing events in rodent striatum [14], while targeted reduction of FUS through similar means affected 374 splicing events [36]. Depending on where TDP43 and FUS bind in relation to the exon-intron boundaries, exon inclusion may be either enhanced or reduced by deficiencies in TDP43 or FUS [13,14,36], pointing toward complex patterns of splicing regulation, and partially explaining the pleiotropic effects of depletion, accumulation, or mislocalization of each protein.…”