Putative RNA-directed adaptive mutations in cancer evolution

Abstract

Understanding the molecular mechanisms behind the capacity of cancer cells to adapt to the tumor microenvironment and to anticancer therapies is a major challenge. In this context, cancer is believed to be an evolutionary process where random mutations and the selection process shape the mutational pattern and phenotype of cancer cells. This article challenges the notion of randomness of some cancer-associated mutations by describing molecular mechanisms involving stress-mediated biogenesis of mRNA-derived small RNAs able to target and increase the local mutation rate of the genomic loci they originate from. It is proposed that the probability of some mutations at specific loci could be increased in a stress-specific and RNA-depending manner. This would increase the probability of generating mutations that could alleviate stress situations, such as those triggered by anticancer drugs. Such a mechanism is made possible because tumor- and anticancer drug-associated stress situations trigger both cellular reprogramming and inflammation, which leads cancer cells to express molecular tools allowing them to "attack" and mutate their own genome in an RNA-directed manner.

Keywords: adaptive mutations; cancer; evolution; mutations; small RNAs.

Figure 1.

(A) By altering the biochemical properties of targeted proteins during translation, a molecular stressor causes the inhibition of translation of the corresponding mRNAs. Stress-induced translationally-stalled mRNAs are cleaved by stress-induced endoribonucleases (step 1). The mRNA fragments are next used as substrates for the biogenesis of small RNAs (step 2). The mRNA-derived small RNAs target the genomic region corresponding to the mRNA precursors and enhance the recruitment of proteins modulating local mutation rate in a direct- or indirect-manner (step 3).

Figure 2.

(A) The subcellular location of proteins (colored circles) rely on the subcellular targeting of their coding mRNAs and subsequent on-site mRNA translation or local translation. The nascent proteins are in different subcellular environment, which may impact on subsequent modifications and interactions with different partners. (B) Co-translational modifications rely on the binding of the protein-modifying enzymes to the translating ribosomes (black circle), or on local translation that positions the nascent proteins in the proximity of specific modifying enzymes (orange circle), or on the binding of protein modifying enzymes to the mRNA 3′-UTR (red circle). The binding of a protein (yellow circle) to mRNA 3′UTR may increase the probability of its interaction with the newly-synthetized protein (blue circle). (C) A stressor affecting a nascent polypeptide chain impact on the mRNA undergoing translation by inhibiting its translation, thus inducing its storage, cleavage, or degradation.

Figure 3.

(A) The cleavage of “precursor” RNAs (primary transcripts) is required for the production of mature RNAs with biological functions. “Mature” RNAs are also cleaved and generate intermediate fragments that are themselves cleaved and modified to generate “derived-small RNAs” having cell regulatory functions. (B) Retrotransposon sequences are transcribed from both DNA strands. Precursor sense RNAs are cleaved by Zucchini. Intermediate RNA fragments are next loaded into Piwi proteins and trimmed by an exoribonuclease up to the regions where Piwi proteins protect small RNA regions from degradation. The resulting sense piRNAs then hybridize to antisense transcripts and induce their cleavage by some Piwi proteins, generating antisense piRNAs that in turn target sense transcripts, into the so-called “ping-pong” process. Several proteins (on the right) involved in piRNA biogenesis are (over)-expressed in cancer cells; protein names followed by * are classified as Cancer/Testis Antigens. (C) A stressor affecting a nascent polypeptide chain impacts on the mRNA undergoing translation by inhibiting its translation and inducing its cleavage. The generated mRNA fragments are then used to produce mRNA-derived small RNAs.

Figure 4.

(A) RNAs produced from intergenic, promoter, or intragenic regions directly and locally recruit proteins (in yellow) involved in chromatin or DNA modifications. (B) Small RNAs form Watson–Crick base-pairing with complementary nascent RNAs leading to the formation on specific loci of complexes enhancing the recruitment of proteins (in yellow) involved in chromatin or DNA modifications. (C) RNA:dsDNA triple-helices (or triplexes) are formed by sequence-specific binding rules where a single-stranded RNA binds in the major groove of the targeted dsDNA by Hoogsteen hydrogen bonding between a purine-rich strand of dsDNA and either a pyrimidine-rich or a purine-rich ssRNA strand. By this mechanism, RNAs direct proteins (in yellow) involved in chromatin or DNA modifications at specific loci. (D) R-loops result from the Watson–Crick base-pairing of an RNA molecule to the cDNA strand displacing the second DNA strand in a single-stranded conformation. (E) RNAs target specific genomic loci by several mechanisms and induce local chromatin and DNA modifications with potential consequences for the local accessibility of mutators and enzymes involved in DNA metabolism. RNAs also induce the formation of structures (e.g., R-loops) that induce ssDNA formation and double-stranded DNA breaks, both of which increase the local mutational rate. Finally, RNAs guide, DNA endonucleases and editing enzymes to targeted loci. Therefore, RNAs could increase the local probability of mutations (RNA-directed mutations) by inducing chromatin modifications (e.g., compaction), DNA modifications (e.g., methylation, ssDNA formation), DNA injuries (e.g., dsDNA breaks), DNA error-prone repair mechanisms, or recruitment of DNA endonucleases and editing enzymes.

Figure 5.

(A) Sustained stress situations trigger chromatin remodeling and the expression of retrotransposons, increasing the transcriptome diversity by impacting for example on antisense transcription. Gene expression reprogramming induced by sustained stresses and retrotransposons can lead to the expression of stem cell-restricted factors and Cancer/Testis Antigens (CTAs), some of which participate in small RNA biogenesis pathways. Antisense transcription of coding genes and of pseudogenes combined with the expression of small RNA biogenesis factors would contribute to the biogenesis of mRNA-derived small RNAs. (B) Tumor cells are in an inflammatory microenvironment due to the release or cellular secretion of DAMP molecules from dead, dying, stressed, or senescent cells. This inflammatory microenvironment mimics a virus infection, which triggers the cell-autonomous innate immune response. (C) In a stressed cell (donor cell), endoribonucleases (e.g., RNase L, IRE1) cleave stress-associated mRNAs in stress granules or on the endoplasmic reticulum. In the context of the tumor inflammatory microenvironment, mRNA fragments may trigger the cellular innate immune response through the activation of receptors like RIG-I and may be released or secreted into the extracellular space by multi-vesicular bodies (MVBs) and extracellular vesicles or by autophagy. A stressed “recipient cell” can capture extracellular RNAs, which (in the context of the tumor inflammatory microenvironment) may be “mistaken” for virus RNAs. The confusion of captured exogenous RNAs for virus RNAs (non-self RNAs), as well as endogenous antisense transcripts produced by the recipient cancer cell, could license mRNA-derived small RNAs to target the corresponding genomic loci for editing.

Figure 6.

(A) Stressor can affect a nascent polypeptide chain and trigger translation inhibition, which can momentarily limit the synthesis of proteins in an unfavorable local environment (“co-translational quality control,” “escape 1”). However, if the stress persists in inducing, for example, an unfolded protein response (ER stress) and if translationally-stalled mRNAs accumulate, they may form stress granules activating a cellular stress response that can collectively either alleviate the stress situation or induce cellular senescence or apoptosis (“Cellular stress response,” “escape 2”). Sustained-stress situations within a cell population induce cell reprogramming (dedifferentiation), leading in particular to retrotransposon expression and antisense transcription. In these conditions, stress-associated mRNAs could be used as substrates to generate mRNA-derived small RNAs through either the “piRNA” or the “endo-siRNA” pathways. Antisense small RNAs could trigger the degradation of the corresponding mRNAs, thereby providing a post-transcriptional adaptive regulatory loop for limiting the accumulation of translationally-stalled mRNAs (PTGS, “escape 3”). mRNA-derived small RNAs may also target the corresponding genomic loci and induce chromatin and DNA modifications leading to transcriptional gene silencing, thereby providing a transcriptional adaptive regulatory loop for limiting the synthesis of stress-associated mRNAs (TGS, “escape 3”). In the context of the virus infection-like microenvironment, which induces the expression of DNA metabolic enzymes that contributing to “genomic plasticity,” including DNA-editing enzymes, small RNAs may increase the local mutational rate of the corresponding and targeted genomic loci through different mechanisms, which would increase the probability of mutation appearance contributing to alleviate the stress situation (escape 4).