1
RNA Regulation in Apoptosis

Christopher von Roretz and Imed-Eddine Gallouzi

McGill University, Department of Biochemistry and Rosalind and Morris Goodman Cancer Research Centre, McIntyre Medical Building Room 915B, 3655 Promenade Sir William Osler, Montreal, Quebec, H3G 1Y6 Canada

1 Balancing Life and Death

2 The Coordinated Process of Apoptosis

2.1 Caspase Cascade

2.2 Extrinsic Activation of Caspases

2.3 Intrinsic Activation of Caspases

2.3.1 Mitochondrial Release of Cytochrome c

2.3.2 Activation of the Apoptosome

2.4 Caspase Substrates

3 Protein Regulators of Apoptosis

3.1 Regulators of Cytochrome c Release

3.1.1 Bcl-2 Family Proteins

3.1.2 BH3-Only Proteins

3.2 Targeting Caspase Activation

3.2.1 DISC Inhibition

3.2.2 Regulation of Apoptosome Activity

3.3 Caspase Inhibitor of Apoptosis Proteins

3.3.1 IAPs

3.3.2 Inhibitors of IAPs

3.4 Post-Translational Regulation of Apoptotic Factors

4 Transcriptional Regulation of Apoptosis

4.1 p53 and p73

4.2 E2F Family

4.3 FOXO Family

4.4 Additional Transcriptional Regulators of Apoptosis

5 Post-Transcriptional Regulation of Apoptosis

5.1 Splicing

5.1.1 Splicing to Regulate Cytochrome c Release

5.1.2 Splicing-Mediated Regulation of Caspases

5.1.3 Nuclear Export of mRNA

5.2 Translation

5.2.1 cis-Elements and trans-Acting Factors that Regulate Translation

5.2.2 HuR as a Regulator of Translation

5.2.3 IRES-Mediated Translation and Apoptosis

5.2.4 microRNAs as Regulators of the Translation of Apoptotic Factors

5.2.5 miRNAs as Inhibitors of Apoptosis

5.2.6 Pro-Apoptotic Roles of miRNA

5.3 mRNA Turnover

5.3.1 The Destabilizing A/U Rich Element

5.3.2 ARE-Mediated Turnover of Apoptosis-Related mRNAs

5.3.3 Alternate Destabilizing Elements

5.3.4 Destabilizing miRNAs

6 Discussion

6.1 Rearranging Deckchairs on the Titanic?

6.2 All a Matter of Time

Acknowledgments

References

Keywords

Alternative splicing

Splicing of pre-mRNA to generate mature mRNA involves the removal of noncoding segments of messenger RNA known as introns. The factors involved in this process determine if certain splice sites are skipped or not, however, yielding different splice variants.

Bcl-2 family proteins

The B-cell lymphoma 2 (Bcl-2) family of proteins all contain one or more Bcl-2 homology (BH) domains. Members of the Bcl-2 family may be either pro- or anti-apoptotic, as they influence mitochondrial outer membrane permeabilization, leading to the release of cytochrome c into the cytoplasm.

Caspase

Cysteine-dependent, aspartic acid-specific proteases (caspases) are generally considered the effectors of apoptotic cell death, cleaving a variety of substrates in order to lead to the organized death of a cell.

Cytochrome c

Cytochrome c (cyt c) is a small polypeptide that plays a role in the electron transport chain as an electron carrier under normal conditions. When released from the mitochondria following mitochondrial outer membrane permeabilization, cytochrome c binding to Apaf-1 protein enables structural changes which ultimately permit the activation of caspase-9.

IRES-mediated translation

Internal ribosomal entry site (IRES)-mediated translation is the process whereby mRNA translation is initiated through the recruitment of translation factors, and consequently, ribosomes, to the mRNA independently of the m7G 5′ cap.

miRNA

microRNA (miRNA) is a single-stranded RNA of approximately 22 nt that can induce the inhibition of translation or decay of a target mRNA upon binding to a mRNA for which a specific miRNA maintains a certain degree of complementarity.

mRNA turnover

The turnover of messenger RNA is the stability of a particular mRNA, which can be modulated by various factors that promote either mRNA decay or stabilization.

Post-transcriptional regulation

Subsequent to the transcription of genetic material to generate messenger RNA, various mechanisms including splicing, localization, translation, and mRNA turnover, can influence the expression of the gene encoded by a target mRNA.

The organized process of apoptotic cell death is tightly regulated, with many protein factors promoting or inhibiting the activity of its key players. A growing number of studies have shown that these numerous regulators of apoptosis are themselves regulated at the level of RNA. Through transcription, the levels of mRNA encoding these factors can be increased or decreased. Less studied is the post-transcriptional regulation of expression for these pro- and anti-apoptotic players. Alternative splicing, effects on translation, and the modulation of mRNA turnover can all influence protein levels of the broad cast of factors involved in apoptosis. While most of the studies delineating these mechanisms have not examined explicitly the RNA regulation of these factors during apoptosis, a small collection of data does suggest that post-transcriptional regulation of apoptotic modulators occurs during the cell death process, thus hinting at a previously underappreciated role for RNA regulation in apoptosis.

1 Balancing Life and Death

All cellular processes depend on balance. Metabolism, motility, and growth all depend on juggling factors that promote or perturb each of these events. Perhaps the most significant of cellular processes, however, is that of cell death when the decision is made – either by the victim cell or by its environment – that life should cease and one, possibly final, balance is tilted either to seal the fate of this cell through death, or to allow it to survive despite assaulting conditions by activating survival mechanisms. At a cellular level, with numerous factors regulating each and every pathway, balance is a matter of life or death.

Several types of cell death exist [1], and while these were initially classified based on their degree of organization, during recent years such a broad-stroke classification has been challenged [1, 2]. While autophagic cell death and necrosis/necroptosis have gained attention more recently [1, 3–7], the best known form of cell death is apoptosis. Derived from the Greek word for “falling off,” as with the leaves from a tree [8], apoptosis has been the most well-studied class of cell death, and is typically considered the default method of ending the life of a cell. In essence, apoptosis is an organized cell death whereby specific processes take place to coordinate the destruction of a cell in a practical manner, to avoid harm to neighboring cells. In fact, given the individualized focus of this process, apoptosis is often referred to as “cell suicide” [3], even though it can be triggered both internally by the victim cell and by external stimuli from neighboring cells or the surrounding environment. Numerous pathways are activated during apoptosis (see below) which integrate many pro- and anti-apoptotic factors to weigh in on the decision to survive or to engage death in response to a stress stimulus. While the activity of these factors has often been studied at great lengths, over the past few years evidence has been mounting to suggest that the expression of apoptotic factors is also an important determinant of apoptotic cell death. Transcription to produce mRNAs encoding these factors has been studied to significant lengths. Conversely, there have been limited attempts at understanding the post-transcriptional mechanisms that influence the expression of apoptotic factors. Of these studies, only a fraction has specifically examined how these regulatory mechanisms may directly influence the progression of apoptosis. In this chapter, attention will be focused on outlining the link between RNA and apoptosis, and a summary provided of how transcription – and, more specifically, post-transcriptional events – may contribute to apoptotic cell death.

2 The Coordinated Process of Apoptosis

Apoptosis will not occur spontaneously, and generally begins with a trigger. The diversity of sources that can activate apoptosis is broad, and ranges from chemical activators, endocrine signaling from neighboring cells, or internal sensors of cell damage [9–13]. Typically, the activation of apoptosis is divided into two categories: (i) that which is triggered externally, known as extrinsic-induced apoptosis; and (ii) that which results from internal signals, termed intrinsic-induced apoptosis [14]. Each of these involves a different sequence of activated factors, although the end goal is the same – to enable the activation of a specific family of proteases known as caspases (Fig. 1).

Fig. 1 Extrinsic and intrinsic pathways of apoptosis. In the extrinsic pathway of apoptotic activation, binding of a death ligand (e.g., Fas ligand, TRAIL) to a death receptor (DR5, Fas) triggers the recruitment of an adaptor protein (FADD, DAXX) and an initiator caspase such as...