Pervasive transcription of the human genome generates RNAs whose mode of formation and functions are largely uncharacterized. Here, we combine RNA-seq with detailed mechanistic studies to describe a ...transcript type derived from protein-coding genes. The resulting RNAs, which we call DoGs for downstream of gene containing transcripts, possess long non-coding regions (often >45 kb) and remain chromatin bound. DoGs are inducible by osmotic stress through an IP3 receptor signaling-dependent pathway, indicating active regulation. DoG levels are increased by decreased termination of the upstream transcript, a previously undescribed mechanism for rapid transcript induction. Relative depletion of polyA signals in DoG regions correlates with increased levels of DoGs after osmotic stress. We detect DoG transcription in several human cell lines and provide evidence for thousands of DoGs genome wide.
Display omitted
•DoGs are a new transcript type generated by readthrough transcription•DoGs are chromatin-bound transcripts highly inducible by osmotic stress•DoGs are induced downstream of more than 10% of human protein-coding genes•DoG induction is mediated through IP3 receptor signaling
Vilborg et al. describe stress-induced transcripts generated by readthrough downstream of protein-coding genes (DoGs). DoGs are regulated by IP3 receptor signaling and remain chromatin-bound. Being long (often >45 kb) and diverse (>2,000 species), DoGs contribute significantly to the human transcriptome.
Stress-induced cleavage of transfer RNAs (tRNAs) into tRNA-derived fragments (tRFs) occurs across organisms from yeast to humans; yet, its mechanistic underpinnings and pathological consequences ...remain poorly defined. Small RNA profiling revealed increased abundance of a cysteine tRNA fragment (5'-tRF
) during breast cancer metastatic progression. 5'-tRF
was required for efficient breast cancer metastatic lung colonization and cancer cell survival. We identified Nucleolin as the direct binding partner of 5'-tRF
. 5'-tRF
promoted the oligomerization of Nucleolin and its bound metabolic transcripts Mthfd1l and Pafah1b1 into a higher-order transcript stabilizing ribonucleoprotein complex, which protected these transcripts from exonucleolytic degradation. Consistent with this, Mthfd1l and Pafah1b1 mediated pro-metastatic and metabolic effects downstream of 5'-tRF
-impacting folate, one-carbon, and phosphatidylcholine metabolism. Our findings reveal that a tRF can promote oligomerization of an RNA-binding protein into a transcript stabilizing ribonucleoprotein complex, thereby driving specific metabolic pathways underlying cancer progression.
The MUSASHI (MSI) family of RNA binding proteins (MSI1 and MSI2) contribute to a wide spectrum of cancers including acute myeloid leukemia. We find that the small molecule Ro 08-2750 (Ro) binds ...directly and selectively to MSI2 and competes for its RNA binding in biochemical assays. Ro treatment in mouse and human myeloid leukemia cells results in an increase in differentiation and apoptosis, inhibition of known MSI-targets, and a shared global gene expression signature similar to shRNA depletion of MSI2. Ro demonstrates in vivo inhibition of c-MYC and reduces disease burden in a murine AML leukemia model. Thus, we identify a small molecule that targets MSI's oncogenic activity. Our study provides a framework for targeting RNA binding proteins in cancer.
Tumourigenesis and cancer progression require enhanced global protein translation
. Such enhanced translation is caused by oncogenic and tumour-suppressive events that drive the synthesis and ...activity of translational machinery
. Here we report the surprising observation that leucyl-tRNA synthetase (LARS) becomes repressed during mammary cell transformation and in human breast cancer. Monoallelic genetic deletion of LARS in mouse mammary glands enhanced breast cancer tumour formation and proliferation. LARS repression reduced the abundance of select leucine tRNA isoacceptors, leading to impaired leucine codon-dependent translation of growth suppressive genes, including epithelial membrane protein 3 (EMP3) and gamma-glutamyltransferase 5 (GGT5). Our findings uncover a tumour-suppressive tRNA synthetase and reveal that dynamic repression of a specific tRNA synthetase-along with its downstream cognate tRNAs-elicits a downstream codon-biased translational gene network response that enhances breast tumour formation and growth.
Activating transcription factor 4 (ATF4) is a master transcriptional regulator of the integrated stress response (ISR) that enables cell survival under nutrient stress. The mechanisms by which ATF4 ...couples metabolic stresses to specific transcriptional outputs remain unknown. Using functional genomics, we identified transcription factors that regulate the responses to distinct amino acid deprivation conditions. While ATF4 is universally required under amino acid starvation, our screens yielded a transcription factor, Zinc Finger and BTB domain-containing protein 1 (ZBTB1), as uniquely essential under asparagine deprivation. ZBTB1 knockout cells are unable to synthesize asparagine due to reduced expression of asparagine synthetase (ASNS), the enzyme responsible for asparagine synthesis. Mechanistically, ZBTB1 binds to the ASNS promoter and promotes ASNS transcription. Finally, loss of ZBTB1 sensitizes therapy-resistant T cell leukemia cells to L-asparaginase, a chemotherapeutic that depletes serum asparagine. Our work reveals a critical regulator of the nutrient stress response that may be of therapeutic value.
Display omitted
•Genetic screens identify transcription factors that regulate amino acid response pathway•ZBTB1 is necessary for the leukemic cell response to asparagine deprivation•ZBTB1 directly binds to ASNS promoter and promotes its transcription•Loss of ZBTB1 sensitizes T cell leukemia cells to L-asparaginase in vivo
Williams et al. identify mediators of the amino acid response pathway in leukemia and demonstrate that ZBTB1 is necessary for cellular proliferation specifically under asparagine deprivation. ZBTB1 associates with the promoter of ASNS and promotes the transcriptional upregulation of ASNS under asparagine deprivation. Loss of ZBTB1 sensitizes leukemic cells to L-asparaginase treatment in vitro and in vivo.
Eukaryotic transfer RNAs can become selectively fragmented upon various stresses, generating tRNA‐derived small RNA fragments. Such fragmentation has been reported to impact a small fraction of the ...tRNA pool and thus presumed to not directly impact translation. We report that oxidative stress can rapidly generate tyrosine‐tRNAGUA fragments in human cells—causing significant depletion of the precursor tRNA. Tyrosine‐tRNAGUA depletion impaired translation of growth and metabolic genes enriched in cognate tyrosine codons. Depletion of tyrosine tRNAGUA or its translationally regulated targets USP3 and SCD repressed proliferation—revealing a dedicated tRNA‐regulated growth‐suppressive pathway for oxidative stress response. Tyrosine fragments are generated in a DIS3L2 exoribonuclease‐dependent manner and inhibit hnRNPA1‐mediated transcript destabilization. Moreover, tyrosine fragmentation is conserved in C. elegans. Thus, tRNA fragmentation can coordinately generate trans‐acting small RNAs and functionally deplete a tRNA. Our findings reveal the existence of an underlying adaptive codon‐based regulatory response inherent to the genetic code.
Synopsis
Whether tRNAs affect gene regulation beyond their central role as adaptors in mRNA translation remains poorly understood. This study uncovers a non‐canonical function for fragments of tyrosine‐coding tRNAGUA in controlling expression of growth promoting genes upon stress.
Oxidative stress‐induced tRNA fragmentation depletes both precursor and mature tRNATyrGUA in human cell lines.
tRNATyrGUA depletion impairs translation of growth genes that are enriched for its cognate tyrosine codons.
Tyrosine‐tRNA fragments are generated in a DIS3L2 ribonuclease‐dependent manner and inhibit RNA‐binding protein hnRNPA1.
Precursor tRNAGUA fragmentation response is conserved in C. elegans.
Ribonuclease‐generated fragments of one particular tRNA control expression of growth‐promoting genes upon stress.
Abstract
Background and aims
Diabetic kidney disease (DKD) is associated with lipid derangements that worsen kidney function and enhance cardiovascular (CVD) risk. The management of dyslipidemia, ...hypertension and other traditional risk factors does not completely prevent CVD complications, bringing up the participation of nontraditional risk factors such as advanced glycation end products (AGEs), carbamoylation and changes in the HDL proteome and functionality. The HDL composition, proteome, chemical modification and functionality were analyzed in nondialysis subjects with DKD categorized according to the estimated glomerular filtration rate (eGFR) and urinary albumin excretion rate (AER).
Methods
Individuals with DKD were divided into eGFR> 60 mL/min/1.73 m
2
plus AER stages A1 and A2 (
n
= 10) and eGFR< 60 plus A3 (
n
= 25) and matched by age with control subjects (eGFR> 60;
n
= 8).
Results
Targeted proteomic analyses quantified 28 proteins associated with HDL in all groups, although only 2 were more highly expressed in the eGFR< 60 + A3 group than in the controls: apolipoprotein D (apoD) and apoA-IV. HDL from the eGFR< 60 + A3 group presented higher levels of total AGEs (20%), pentosidine (6.3%) and carbamoylation (4.2 x) and a reduced ability to remove
14
C-cholesterol from macrophages (33%) in comparison to HDL from controls. The antioxidant role of HDL (lag time for LDL oxidation) was similar among groups, but HDL from the eGFR< 60 + A3 group presented a greater ability to inhibit the secretion of IL-6 and TNF-alpha (95%) in LPS-elicited macrophages in comparison to the control group.
Conclusion
The increase in apoD and apoA-IV could contribute to counteracting the HDL chemical modification by AGEs and carbamoylation, which contributes to HDL loss of function in well-established DKD.
The human genome contains 61 codons encoding 20 amino acids. Synonymous codons representing a given amino acid are decoded by a set of transfer RNAs (tRNAs) called isoacceptors. We report the ...surprising observation that two isoacceptor tRNAs that decode synonymous codons become modulated in opposing directions during breast cancer progression. Specifically, tRNA
became upregulated, whereas tRNA
became repressed as breast cancer cells attained enhanced metastatic capacity. Functionally, tRNA
promoted and tRNA
suppressed metastatic colonization in mouse xenograft models. These tRNAs mediated opposing effects on codon-dependent translation of growth-promoting genes, consistent with genomic enrichment or depletion of their cognate codons in mitotic genes. Our findings uncover a specific isoacceptor tRNA pair that act in opposition, divergently impacting growth-regulating genes and a disease phenotype. Degeneracy of the genetic code can thus be biologically exploited by human cancer cells via tRNA isoacceptor shifts that causally facilitate the transition toward a growth-promoting state.
PDF
