Rapid Nucleolytic Degradation of the Small Cytoplasmic Y RNAs during Apoptosis

Abstract

We have investigated the fate of the RNA components of small ribonucleoprotein particles in apoptotic cells. We show that the cytoplasmic Ro ribonucleoprotein-associated Y RNAs are specifically and rapidly degraded during apoptosis via a caspase-dependent mechanism. This is the first study describing the selective degradation of a specific class of small structural RNA molecules in apoptotic cells. Cleavage and subsequent truncation of Y RNAs was observed upon exposure of cells to a variety of apoptotic stimuli and were found to be inhibited by Bcl-2, zinc, and several caspase inhibitors. These results indicate that apoptotic degradation of Y RNAs is dependent on caspase activation, which suggests that the nucleolytic activity responsible for hY RNA degradation is activated downstream of the caspase cascade. The Y RNA degradation products remain bound by the Ro60 protein and in part also by the La protein, the only two proteins known to be stably associated with intact Ro ribonucleoprotein particles. The size of the Y RNA degradation products is consistent with the protection from degradation of the most highly conserved region of the Y RNAs by the bound Ro60 and La proteins. Our results indicate that the rapid abrogation of the yet unknown function of Y RNAs might be an early step in the systemic deactivation of the dying cell.Apoptosis is a form of cell death characterized by distinct morphological and biochemical alterations. Morphologically, apoptotic cells are characterized by chromatin condensation, cell shrinkage, fragmentation of the nucleus, and partition of cytoplasm and nucleus into membrane bound-vesicles (apoptotic bodies) (1). During the last 5 years, many of the molecules that participate in the biochemical pathway mediating apoptosis have been identified. A major role in this pathway is played by caspases, cysteine proteases with aspartic acid substrate specificity (2). Proteins cleaved by caspases appear to be structural proteins essential for maintaining nuclear and cytoplasmic architecture and enzymes essential for repair of damaged cell components (reviewed in Ref.3). A prominent nuclear event during apoptosis is internucleosomal cleavage of DNA, recognized as a "DNA ladder" on conventional agarose gel electrophoresis (4). The endonuclease activity responsible for apoptotic degradation of chromosomal DNA has recently been identified (5). The activity depends on two interacting proteins, one of which contains the endonuclease activity (caspase-activated deoxyribonuclease (CAD) 1 ), which is retained in the cytoplasm in an inactive form due to its association with the second protein (inhibitor of CAD). Caspase activation in apoptotic cells leads to cleavage of the inhibitor of CAD, thereby releasing active CAD resulting in DNA fragmentation in the nuclei (5, 6).Much less is known about cleavage and degradation of RNA in apoptotic cells. An increased rate of mRNA turnover has been suggested (7, 8) as well as mitochondrial 16 S ribosomal RNA degradation (9), but no nuclease associated with specif...

Physical interaction of apoptosis-inducing factor with DNA and RNA

“…This result was obtained using two distinct methods for the evaluation of total cellular RNA content, namely RNA extraction followed by horizontal agarose gel electrophoresis (Figure 4b) or staining with the RNA probe pyronin Y (Crissman et al, 1985;Halicka et al, 2000), followed by cytofluorometric evaluation of the per-cell RNA content (Figure 4c). Thus, in accord with the literature (Rutjes et al, 1999;King et al, 2000;Asselbergs and Widmer, 2003), apoptotic RNA degradation is caspase-dependent, and, as shown in Figure 3b and c, AIF-independent.…”

Circulating microRNA signature of genotype‐by‐age interactions in the long‐lived Ames dwarf mouse

“…Recently, deep sequencing unraveled that tRNA, rRNA, snoRNA, and YRNA can undergo processing into smaller RNA molecules (Rutjes et al ., 1999; Rother & Meister, 2011; Sobala & Hutvagner, 2011). These new small RNAs are being noticed only now because most studies focused on sequencing reads derived from miRNAs.…”

“…These new small RNAs are being noticed only now because most studies focused on sequencing reads derived from miRNAs. Accumulating evidence suggests that these RNA fragments are functional in physiological and pathological conditions (Rutjes et al ., 1999; Haussecker et al ., 2010; Anderson & Ivanov, 2014). In particular, tRNA halves have been described in the cytoplasm of stressed cultured cells where they have been reported to function in key biological processes, that is, translation and apoptosis (Ivanov et al ., 2011; Anderson & Ivanov, 2014) and may be involved in human diseases including cancer, neurodegeneration, and infection [reviewed in Anderson & Ivanov (2014)].…”

“…More recently, deep sequencing technologies allowed the identification of new types of small RNAs derived from the processing of already known sncRNAs including tRNA (Rutjes et al ., 1999; Rother & Meister, 2011; Sobala & Hutvagner, 2011). There is evidence implicating these derivatives of sncRNAs in cell‐to‐cell communication both in normal biology and in disease states (Cortez et al ., 2011; Hoy & Buck, 2012; Shah & Calin, 2012; Turchinovich et al ., 2012; Kosaka et al ., 2013).…”

Rearrangement of nuclear ribonucleoprotein (RNP)‐containing structures during apoptosis and transcriptional arrest

“…Rutjes et al (1999) suggest that Y RNAs are nucleolytically degraded during apoptosis and the consequent block of Y RNA function could represent an early step of apoptotic cell death. (Nadano and Sato 2000) proved that RNA degradation may be involved in the inhibition of protein synthesis during apoptosis initiated by death receptor engagement, and according to (Degen et al (2000) the mechanism of rRNA cleavage can be considered as a non-specific event reflecting the generalized shut-down of translation.…”