Heterogeneous Nuclear Ribonucleoproteins C1 and C2 Associate with the RNA Component of Human Telomerase

Abstract

Here we demonstrate that heterogeneous nuclear ribonucleoproteins (hnRNPs) C1 and C2 can associate directly with the integral RNA component of mammalian telomerase. The binding site for hnRNPs C1 and C2 maps to a 6-base uridylate tract located directly 5 to the template region in the human telomerase RNA (TR) and a 4-base uridylate tract directly 3 to the template in the mouse TR. Telomerase activity is precipitated with antibodies specific to hnRNPs C1 and C2 from cells expressing wild-type human TR but not a variant of the human TR lacking the hnRNPs C1 and C2 binding site, indicating that hnRNPs C1 and C2 require the 6-base uridylate tract within the human TR to associate with the telomerase holoenzyme. In addition, we demonstrate that binding of hnRNPs C1 and C2 to telomerase correlates with the ability of telomerase to access the telomere. Although correlative, these data do suggest that the binding of hnRNPs C1 and C2 to telomerase may be important for the ability of telomerase to function on telomeres. The C proteins of the hnRNP particle are also capable of colocalizing with telomere binding proteins, suggesting that the C proteins may associate with telomeres in vivo. Therefore, human telomerase is capable of associating with core members of the hnRNP family of RNA binding proteins through a direct and sequence-specific interaction with the human TR. This is also the first account describing the precise mapping of a sequence in the human TR that is required to associate with an auxiliary component of the human telomerase holoenzyme.Telomeres have many functions, including organizing chromosomes during meiosis, protecting chromosome ends from nucleolytic digestion and end-to-end fusion events, and facilitating complete replication of the chromosomes (3, 22, 64). During normal lagging strand replication of linear chromosomes, the ends are left with a gap between the final RNA priming event and the terminus (48,63). In the absence of any compensatory mechanism, telomere length decreases with each cell division cycle until the cell reaches a state of proliferative failure. Thus, telomeres also serve as a source of expendable DNA, avoiding the loss of coding DNA sequences.Telomerase is a cellular ribonucleoprotein reverse transcriptase that can maintain the length of telomeres by using its integral RNA component as a template for the addition of TTAGGG repeats onto the 3Ј end of linear chromosomes.

In vivo identification of ribonucleoprotein-RNA interactions

“…4Cc is a positive control showing that phosphocAMP response element-binding protein is observed in the cells after BDNF treatment. Further, hnRNP A1, hnRNP C1͞C2, and ribosomal protein L22 were found to associate with telomerase mRNA (44)(45)(46) and have also been found to increase under BDNF-stimulated conditions. Because telomerase dysfunction has been associated with the onset of rheumatoid arthritis (47), and also because the same RBPs bind to ank RNA, both of which are associated with arthritic physiology, it is tempting to speculate that these RBPs may be involved in coordinating aspects of normal bone physiology.…”

TERRA transcripts are bound by a complex array of RNA-binding proteins

“…Interestingly, some of the identifi ed RBPs have been previously reported to participate in regulation of telomeres and telomerase, as mentioned above. In particular, hnRNP A1, A2B1 and D / AUF1 bind to telomeres 11 -13 (reviewed in Ford et al 14 ) and hnRNP A1, D and GAR1 bind to telomerase 12,15,16,22 . Th e second approach allowed the identifi cation of those RBPs that bind more abundantly to the biotinylated TERRA ( Fig.…”

Analysis of PTEN Complex Assembly and Identification of Heterogeneous Nuclear Ribonucleoprotein C as a Component of the PTEN-associated Complex

“…It is a compo- nent of the spliceosome complex (38), and a recent study using a hnRNP C knockdown approach identified BCL2L12 and PCBP4 as genes whose splicing is regulated by hnRNP C (39). Additional functions, such as regulating telomerase activity (40), XIAP translation through the IRES promoter (41), and response to hydrogen peroxide stress (42,43) have been attributed to hnRNP C. The interaction of PTEN with hnRNP C and the complex assembly may be important in regulating these known hnRNP C functions.…”