Identification of Undifferentiated Embryonic Cell Transcription Factor 1 as a Potential Substrate of Carboxyl-Terminal Domain Small Phosphatases

Abstract

Protein phosphorylation and dephosphorylation play important roles in intracellular communication.1 C-terminal domain (CTD) small phosphatase 1 (CTDSP1), previously known as small CTD phosphatase 1 (SCP1), is an enzyme that preferentially dephosphorylates serine residues in the CTD of RNA polymerase II (RNAPII).2 CTDSP1 catalyzes the dephosphorylation of the fifth phosphorylated serine of Y 1 S 2 P 3 T 4 S 5 P 6 S 7 in the RNAPII CTD.3 CTD small phosphatase 2 (CTDSP2), previously known as small CTD phosphatase 2 (SCP2), and CTD small phosphatase like 1 (CTDSPL1), previously known as small CTD phosphatase 3 (SCP3), have a similar sequence and structure to CTDSP1.4 CTD small phosphatases (CTDSPs) have been documented as neuronal development regulators by silencing neuronal genes.5 Recent studies suggest that cell division cycle associated 3 (CdcA3)6 and receptor-regulated SMADs (R-SMADs) 7,8 can be dephosphorylated by CTDSPs, and that CTDSPs are involved in cell cycle regulation and differentiation.Undifferentiated embryonic cell transcription factor 1 (Utf1), which is specifically expressed in two pluripotent cell lines (mouse embryonic carcinoma cells and mouse embryonic stem cells), is a key player in embryonic cell development and cell fate determination.9 Interestingly, Utf1 is a Eutherian-specific pluripotency marker.10 Utf1 is a chromatinassociated protein with repressor activity and is also involved embryonic stem (ES) cell differentiation.11 A recent study proposed that Utf1 prevents excessive inhibition of bivalent genes by blocking polycomb repressive complex 2 (PRC2) binding and subsequent silencing via Histone 3 (H3) lysine 27 trimethylation. The same study also proposes that Utf1 fine-tunes bivalent gene expression by tagging newly transcribed mRNAs in the nucleus for cytoplasmic degradation. Therefore, Utf1 acts as an epigenetic and translationmodulating factor, and contributes to regulation of pluripotency.12 In recent studies, the role of Utf1 in cervical carcinoma and carcinogenesis was defined.13 Two phosphoproteomics studies of human embryonic stem cells reported that Utf1 could be phosphorylated during their differentiation. 14,15Utf1 has five phosphorylated serine and threonine residues: S18, T35, S42, S54, and S245. The roles of these phosphorylations might be related to regulation of Utf1-binding to target proteins or nucleic acids. Additional investigation to reveal the biological roles of these phosphorylations is necessary to understand Utf1's role in ES cell differentiation. Furthermore, studies describing how these phosphorylated residues are dephosphorylated are indispensable to ascertain how Utf1 is regulated.The 3-dimensional structures of CTDSP1 (e.g., PDB ID: 2GHT, 2GHQ, 1TAO), 3,16 CTDSP2 (PDB ID: 2Q5E), 17 and CTDSPL1 (PDB ID: 2HHL) 17 have been solved by X-ray crystallography. The X-ray crystal structures of a dominantnegative form of human CTDSP1 bound to mono-and diphosphorylated peptides (PDB ID: 2GHQ and 2GHT) encompassing the CTD heptad repeat in RNAPII (Y 1 S 2 P 3 T 4 ...