No abstract

The DPF (double PHD finger) domain consists of two PHD fingers organized in tandem. The two PHD-finger domains within a DPF form a single structure that interacts with the modification of the N-terminal histone fragment in a way different from that for single PHD fingers. Several histone modifications interacting with the DPF domain have already been identified. They include acetylation of H3K14 and H3K9, as well as crotonylation of H3K14. These modifications are found predominantly in transcriptionally active chromatin. Proteins containing DPF belong to two classes of protein complexes, which are the transcriptional coactivators involved in the regulation of the chromatin structure. These are the histone acetyltransferase complex belonging to the MYST family and the SWI/SNF chromatin-remodeling complex. The DPF domain is responsible for the specificity of the interactions between these complexes and chromatin. Proteins containing DPF play a crucial role in the activation of the transcription of a number of genes expressed during the development of an organism. These genes are important in the differentiation and malignant transformation of mammalian cells.

Chromatin remodelling multiprotein complexes play an important role in regulation of gene expression in embryogenesis and in the adult organism. Mutations in the subunits of the complexes are often lethal or lead to developmental defects. Complexes consist of core subunits and a specific module. The core consists of ATPase and structure subunits, specific subunits of the module are necessary for chromatin binding. PHF10 (PHD finger protein 10) is a subunit of the PBAF (polybromo-associated BAF) chromatin remodelling complex subfamily. Conserved and highly regulated PHF10 is ubiquitously expressed in mammals as four different isoforms. The isoforms of PHF10 differ by domain structures and posttranslational modifications. All isoforms are highly regulated and included in the PBAF complex in a mutually exclusive manner. Two of the PHF10 isoforms (PHF10-P) are expressed at a high level in neuronal and myeloid progenitors and are necessary for cell proliferation. These isoforms contain PHD (plant homeodomain) fingers for nucleosome binding and recruit RNA polymerase II on the promoters of cell cycle genes. Two other isoforms (PHF10-S) instead of PHD have PDSM (phosphorylation-dependent sumoylation motif), the motif for SUMO1 conjugation. PHF10 is the most unstable subunit of the PBAF complex. Stability can alter the turnover rate of the subunits of the PBAF complex. All PHF10 isoforms are degraded by β-TrCP ubiquitin ligase but PHF10-S isoforms contain a cluster of serins (X-cluster) for multiple phosphorylation by casein kinase I. This phosphorylation protects the β-TrCP degron from β-TrCP recognition and subsequently stabilizes the PHF10-S isoforms. Thus, the incorporation of PHF10 isoforms with different phosphorylation patterns and different stability into the PBAF complexes alters the functions of the entire PBAF complex and determines the range of genes undergoing remodelling.

The mammalian PBAF subfamily of SWI/SNF chromatin remodeling complexes plays a wide role in the regulation of gene expression. PHF10 is a subunit of the signature module of PBAF, responsible for its interaction with chromatin. PHF10 is represented by four different isoforms, which are alternatively incorporated in the complex. Two of PHF10 isoforms lacking C-terminal PHD domains contain a cluster of phosphorylated serine residues, designated as X-cluster. In the present study, we explore the phosphorylation of the X-cluster in detail. We identified additional phosphorylated serine residues and designated them as either frequently or rarely phosphorylated. The X-cluster consists of two independently phosphorylated subclusters. Phosphorylation of the second subcluster depends on phosphorylation of a primary serine 327. These two subclusters surround a sequence, which is predicted to be a nuclear localization sequence (NLS3). The NLS3 does not affect localization of PHF10 isoforms. However, it is essential for X-cluster phosphorylation and increased stability of isoforms that lack PHD. Conversely, the presence of NLS3 signal in isoforms that contain C-terminal PHD domains reduces their stability. Thus, phosphorylation of PHF10 isoforms regulates their cell level, determining the rate of incorporation in PBAF. This may alter the pattern of PBAF regulated genes.

The PBAF chromatin remodeling complex interacts with many transcriptional activators and is recruited to target chromatin regions. PBAF plays an important role in maintaining and modifying the chromatin structure in mammalian cells. A subunit of the PBAF complex, the PHF10 transcription factor, is required for proliferation of neuronal precursors in the early stages of mouse brain development and gene expression in differentiated neurons. We showed that PHF10 interacts with the protein product of the early response gene c-Fos, the c-FOS transcriptional activator, which is expressed in response to the induction of long-term potentiation (LTP). LTP induction triggers the transcription of genes and the synthesis of proteins that provide changes that lead to the establishment of long-term contacts between neurons. We showed that in cells in differentiated neuronal culture, after the induction of LTP, expression of c-FOS, which is initially localized in the cytoplasm and then moves to the nucleus, begins. PHF10 is expressed in neuronal cells prior to LTP induction and has nuclear localization. However, 1 h after LTP induction, PHF10 is detected in the cytoplasm together with c-FOS, and then moves into the nucleus with it. Importantly, this behavior of PHF10 in response to KCl stimulation is specific for neuronal cultures. It is assumed that during LTP, PHF10 together with c-FOS participates in the activation of secondary response genes that regulate the maintenance of plastic modifications and homeostasis of neuronal synapses. The PHF10 export from the nucleus and its rapid return together with c-FOS to the nucleus is possibly necessary for the rapid modulation of expression of target secondary response genes during LTP.