hCLE/C14orf166 Associates with DDX1-HSPC117-FAM98B in a Novel Transcription-Dependent Shuttling RNA-Transporting Complex

Abstract

hCLE/C14orf166 is a nuclear and cytoplasmic protein that interacts with the RNAP II, modulates nuclear RNA metabolism and is present in cytoplasmic RNA granules involved in localized translation. Here we have studied whether hCLE shares common interactors in the nucleus and the cytosol, which could shed light on its participation in the sequential phases of RNA metabolism. Nuclear and cytoplasmic purified hCLE-associated factors were identified and proteins involved in mRNA metabolism, motor-related proteins, cytoskeletal and translation-related factors were found. Purified hCLE complexes also contain RNAs and as expected some hCLE-interacting proteins (DDX1, HSPC117, FAM98B) were found both in the nucleus and the cytoplasm. Moreover, endogenous hCLE fractionates in protein complexes together with DDX1, HSPC117 and FAM98B and silencing of hCLE down-regulates their nuclear and cytosolic accumulation levels. Using a photoactivatable hCLE-GFP protein, nuclear import and export of hCLE was observed indicating that hCLE is a shuttling protein. Interestingly, hCLE nuclear import required active transcription, as did the import of DDX1, HSPC117 and FAM98B proteins. The data indicate that hCLE probably as a complex with DDX1, HSPC117 and FAM98B shuttles between the nucleus and the cytoplasm transporting RNAs suggesting that this complex has a prominent role on nuclear and cytoplasmic RNA fate.

The mammalian tRNA ligase complex mediates splicing of XBP1 mRNA and controls antibody secretion in plasma cells

“…To address whether the subcellular localization of RTCB and archease is compatible with a role in both processes, we performed subcellular fractionation experiments (Fig1D) and immunofluorescence staining (Fig1E) in control cells and upon UPR induction by means of thapsigargin (Tg) treatment, an inhibitor of ER Ca 2+ -ATPases. We detected RTCB both in the nucleus and in the cytoplasm, which is in agreement with a recent report identifying the tRNA ligase as part of RNA transport complexes shuttling between these two compartments (Perez-Gonzalez et al , 2014). In contrast, archease was strongly enriched in the cytoplasm (Fig1D) and found in perinuclear regions stained by the ER membrane marker calnexin (Fig1E).…”

“…Even though tRNA splicing is thought to be a predominantly nuclear process (De Robertis et al , 1981; Nishikura & De Robertis, 1981; Lund & Dahlberg, 1998), we found archease and the majority of RTCB localizing to cytoplasmic compartments. This subcellular distribution of RTCB coincides with a recent report describing the tRNA ligase as part of an RNA transport complex shuttling between nucleus and cytoplasm (Perez-Gonzalez et al , 2014). This flexible localization of RTCB and the cytoplasmic distribution of archease suggest that shuttling of the tRNA ligase not only supports RNA transport but also enables guanylation of RTCB by archease.…”

Proteomic analysis of FUS interacting proteins provides insights into FUS function and its role in ALS

“…Although we did not identify FA98B/FAM98B, a fourth component of the tRNA splicing ligase complex as a FUS interacting partner, we did identify its closest human homolog, FA98A/FAM98A (Supplemental Table 1). The FUS-interacting RTCB/CN166/DDX1 complex was also reported to play a role in nucleo-cytoplasmic RNA shuttling [74], a function shared by FUS [9], suggesting potential functional cooperation. The familial ALS-related FUS mutations cause cytoplasmic mis-localization of FUS [3, 4, 31–34].…”

Expression and clinical significance of C14orf166 in esophageal squamous cell carcinoma

“…Thus, C14orf166 may represent a potential biomarker to guide the selection of therapeutic strategy and improve survival, although those who received adjuvant therapy in the present study could not demonstrate this advantage, which was limited by the small size of the study. Furthermore, C14orf166 has been reported to be detected in the serum of pancreatic carcinoma (13), which indicates a potential novel biomarker for early diagnosis and an aid in therapy design. Therefore, further investigations are required to fully elucidate this potential biomarker.…”

“…It is regarded as a shuttling protein that transports RNAs between the nucleus and cytoplasm, and serves a prominent role in RNA fate and selective gene expression. It has been demonstrated that the C14orf166-DDX1-HSPC117-FAM98B complex helps RNAs shuttle back and forth (13). C14orf166 has also been demonstrated to be a binding partner of Janus kinase (JAK) 2, which can activate excessive signal transducer and activator of transcription (STAT) 3 function to unbalance its tumor promotion and anti-tumor function, hence initiating tumorigenesis (14,15).…”