Conserved interactions of the splicing factor Ntr1/Spp382 with proteins involved in DNA double-strand break repair and telomere metabolism

Abstract

The ligation of DNA double-strand breaks in the process of non-homologous end-joining (NHEJ) is accomplished by a heterodimeric enzyme complex consisting of DNA ligase IV and an associated non-catalytic factor. This DNA ligase also accounts for the fatal joining of unprotected telomere ends. Hence, its activity must be tightly controlled. Here, we describe interactions of the DNA ligase IV-associated proteins Lif1p and XRCC4 of yeast and human with the putatively orthologous G-patch proteins Ntr1p/Spp382p and NTR1/TFIP11 that have recently been implicated in mRNA splicing. These conserved interactions occupy the DNA ligase IV-binding sites of Lif1p and XRCC4, thus preventing the formation of an active enzyme complex. Consistently, an excess of Ntr1p in yeast reduces NHEJ efficiency in a plasmid ligation assay as well as in a chromosomal double-strand break repair (DSBR) assay. Both yeast and human NTR1 also interact with PinX1, another G-patch protein that has dual functions in the regulation of telomerase activity and telomere stability, and in RNA processing. Like PinX1, NTR1 localizes to telomeres and associates with nucleoli in yeast and human cells, suggesting a function in localized control of DSBR.

Spp382p Interacts with Multiple Yeast Splicing Factors, Including Possible Regulators of Prp43 DExD/H-Box Protein Function

“…Spp2p functions with Prp2p in splicing (Silverman et al 2004) while Spp382p and Pxr1p act with Prp43p in splicing and ribosome biogenesis, respectively (Guglielmi and Werner 2002;Tsai et al 2005;Boon et al 2006;Pandit et al 2006;Tanaka et al 2007). Pxr1p and Spp382p also influence telomerase activity and genomic stability (Lin and Blackburn 2004;Herrmann et al 2007), suggesting additional contributions to nuclear function. The biological roles for Sqs1p and the Ylr271W product are unknown but because single (Giaever et al 2002) or combined (S. Pandit and B. C. Rymond, unpublished results) null mutants of these genes are viable, neither one makes a critical contribution to cellular biochemistry under standard conditions.…”

Restricting the ligation step of non-homologous end-joining

“…But what happens if Ntr1 cannot interact with Lif1 and how this may occur? As reported, one of the telomere-associated proteins colocalizing with Ntr1 is Rap1 [23]. Rap1 directly binds the telomeres and the amount bound increases with number of telomeric repeats.…”

“…Recently, Herrmann et al discovered another Lif1-interacting partner, Ntr1 [23], an essential protein that functions in the spliceosome disassembly process [24]. Interestingly, the region in Lif1 responsible for interaction with Ntr1 (residues 220-240) overlaps to a large extent with the region involved in binding to Dnl4 (residues 205-228) [25], indicating that Ntr1 competes with Dnl4 for binding to Lif1 (Fig.…”

“…Outstanding questions are why a cell might need to channel Ntr1 into complex with Lif1 and where this might happen. Given that Ntr1 was demonstrated to co-localize with telomere-associated proteins, it is reasonable to suggest that this protein functions to disrupt the ligase complex at telomeres, causing the down-regulation of NHEJ at these chromosomal regions [23]. But what happens if Ntr1 cannot interact with Lif1 and how this may occur?…”

The G-Patch Domain of Mason-Pfizer Monkey Virus Is a Part of Reverse Transcriptase

“…The GPD is an approximately 40-amino-acid-long region that is characterized by the presence of six highly conserved glycine residues in the sequence hhxxxGaxxGxGhGxxxxG(x) n G, where "h" stands for bulky, hydrophobic residues (I, L, V, or M); "a" stands for aromatic residues (F, Y, or W); and "x" stands for any residue. Various G-patch-containing proteins have been found to be involved in mRNA splicing (14,24,28,32) and DNA repair (7,11) and to be overexpressed in the great majority of breast cancer cases (19). In most G-patch-containing proteins, this conserved domain is usually combined with other well-defined RNA binding domains, such as the RRM, dsRED, SWAP, R3H, C4, and C2H2 fingers (1).…”