DOI 10.1515/hsz-2013-0264 Biol. Chem. 2014; 395(4): 375–386 Review Michael Lienhard Schmitz*, Alfonso Rodriguez-Gil and Juliane Hornung Integration of stress signals by homeodomain interacting protein kinases Abstract: The family of homeodomain interacting protein N-terminal kinase domain and multiple functional kinases (HIPKs) consists of four related kinases, HIPK1 domains in the C-terminus. In contrast to these canonical to HIPK4. These serine/threonine kinases are evolution- HIPKs (HIPK1-3), the family member HIPK4 only shares ary conserved and derive from the yeast kinase Yak1. The the kinase domain with the other family members and largest group of HIPK phosphorylation substrates is repre- thus is very likely to display distinct biological roles. The sented by transcription factors and chromatin-associated HIPK family of protein kinases belongs to the CMGC (con- regulators of gene expression, thus transferring HIPK- taining CDK, MAPK, GSK, CLK families) group of protein derived signals into changes of gene expression programs. kinases (Manning et al., 2002). Within this group, HIPKs The HIPKs mainly function as regulators of developmen- are most closely related to the family of dual-specifity tal processes and as integrators of a wide variety of stress tyrosine phosphorylation regulated kinases (DYRK), as signals. A number of conditions representing precarious schematically shown in Figure 1. Most of the information situations, such as DNA damage, hypoxia, reactive oxygen discussed here has been gathered for HIPK2, which is the intermediates and metabolic stress affect the function of most-studied member of this family of protein kinases. HIPKs. The kinases function as integrators for these stress HIPK2 is detected much more frequently than its sister signals and feed them into many different downstream kinases when unbiased screens such as functional experi- effector pathways that serve to cope with these precarious ments or yeast two-hybrid screens are employed. The situations. HIPKs do not function as essential core com- nature of this bias is not known and the preferred detec- ponents in the different stress signaling pathways, but tion of HIPK2 may suggest its special biological relevance. rather serve as modulators of signal output and as con- The phylogenetic ancestor of HIPKs and DYRKs is the nectors of different stress signaling pathways. Their cen- yeast Yak1 protein, which already shows most of the prin- tral role as signaling hubs with the ability to shape many ciple functional features displayed by the canonical HIPK downstream effector pathways frequently implies them in family members. proliferative diseases such as cancer or fibrosis. Keywords: cell stress; HIPKs; phosphorylation; posttrans- lational modification; signal integration. Yak1, an ancestral kinase linking cell stress to transcriptional *Corresponding author: Michael Lienhard Schmitz, Institute of regulation Biochemistry, Medical Faculty, Friedrichstrasse 24, Justus-Liebig- University, 35392 Giessen, Germany, The Yak1 protein negatively regulates cell proliferation and e-mail: [email protected] its kinase activity is upregulated during S phase arrest of Alfonso Rodriguez-Gil and Juliane Hornung: Institute of Biochemistry, Medical Faculty, Friedrichstrasse 24, Justus-Liebig- yeast cells (Garrett et al., 1991). Also the late mitotic regu- University, 35392 Giessen, Germany latory network controlling cyclin destruction is controlled by Yak1, as revealed by genetic evidence. The growth defect of several late mitotic mutants leading to cell cycle arrest in anaphase could be rescued by overexpression of Evolutionary conservation of the Yak1 (Jaspersen et al., 1998), but the molecular mecha- HIPK family nisms mediating the rescue remain to be established. Besides its involvement in cell cycle regulation, many The members of the evolutionary conserved family studies have identified Yak1 as a stress sensor that medi- of HIPKs have a similar basic architecture, with an ates gene regulation in response to heat shock, acid stress 376 M.L. Schmitz et al.: HIPKs as stress signal integrators DYRK2 Initial evidence for the regulation of Yak1 by glucose came DYRK3 from a study describing that deletion of the YAK1 gene DYRK1A DYRK4 allows growth of yeast cells lacking the catalytic subunit of the cAMP-dependent protein kinase PKA (Garrett and DYRK1B Broach, 1989). Yak1 is not only negatively regulated by PKA, but also by TOR, the second relevant nutrient- sensing kinase pathway (Martin et al., 2004; Schmelzle HIPK3 et al., 2004). In unstressed cells, the Yak1 protein local- HIPK1 izes throughout the entire cell. Induction of stress, by HIPK2 removal of glucose or rapamycin-induced inhibition of HIPK4 TOR, results in nuclear accumulation of Yak1 (Moriya et al., 2001). Cytosolic localization of Yak1 requires PKA- dependent phosphorylation of Yak1 on Ser295 (Lee et al., 2011). Autophosphorylation at Thr335 enables binding of a CMG group yeast 14-3-3 protein, which decreases the catalytic activity of Yak1 and also contributes to the cytoplasmic retention of inactive Yak1 under high glucose conditions (Lee et al., 2011). The group of Yak1-regulated transcription factors Figure 1 The CMG group of kinases in the phylogenetic tree of includes the stress responsive transcription factors heat human kinases (Manning et al., 2002). shock factor protein 1 (Hsf1) and Msn2/Msn4. They are direct phosphorylation targets of Yak1 under conditions where PKA activity is lowered by glucose depletion (Lee and metabolic stress. Glucose availability is reflected at the et al., 2008). While it is known that Yak1-mediated phos- levels of the two kinases PKA (protein kinase A) and TOR phorylation of Hsf1 increases the DNA-binding activity of (target of rapamycin), which both regulate the localization the transcription factor, the functional relevance of Msn2 and activity of Yak1, as schematically shown in Figure 2. phosphorylation remains to be established (Lee et al., 2008). Genetic evidence shows that Msn2/Msn4 trigger the High Low expression of enzymes mediating synthesis of the storage glucose glucose carbohydrate glycogen. In addition, these transcription factors augment Yak1 expression and thus establish an cAMP autoregulatory loop (Smith et al., 1998). Limitation of TOR PKA TOR PKA glucose also leads to Yak1-dependent phosphorylation of Pop2/Caf1, a homolog of the human cNOT7 protein which is contained in the Ccr4-Not (carbon catabolite repressor 4 Yak1 p negative on TATA-less) complex (Moriya et al., 2001). This multi-subunit complex has been implicated in many dif- 14-3-3 ferent aspects of the mRNA life cycle (Miller and Reese, 2012) and interestingly Yak1 seems also to associate with Yak1 other components of this complex, such as cell division Glycogen cycle (Cdc)39/Not1 (http://www.yeastgenome.org/). These p p synthesis findings raise the possibility that Yak1 also contributes to YAK1 Hsf1 Msn2/4 post-transcriptional gene regulation. A further study pro- posed the transcription factor Haa1 as a Yak1 target under conditions of acidic stress (Malcher et al., 2011). The same study also implied the transcription factors Sok2 and Phd1 as downstream targets of Yak1, showing that this kinase Figure 2 Glucose responsiveness of the yeast Yak1 protein. Under can regulate a multitude of transcription factors (Malcher conditions of high glucose, Yak1 is retained in the cytosol by TOR et al., 2011). and PKA-dependent signals. Low glucose levels allow phospho- rylation and activation of Hsf1 and Msn2/4, which in turn lead to Yak1 also acts on transcriptional coregulators such induced synthesis of Yak1 and of enzymes catalyzing glycogen as the corepressor Crf1 which participates in the negative synthesis. regu lation of ribosomal protein genes and thus of ribosome M.L. Schmitz et al.: HIPKs as stress signal integrators 377 biogenesis. TOR/PKA-mediated signals negatively regulate Splice variant S AL NLS1&2 Yak1 and therefore maintain the inhibitory protein Crf1 in -Ubi the cytoplasm. TOR inactivation leads to Yak1 induction, kinase domain Interaction inhibit.S, Q, A HIPK2 which in turn phosphorylates and activates Crf1 (Martin SIM Caspase et al., 2004). The phosphorylated corepressor accumu- S AL NLS1 -Ubi lates in the nucleus and inhibits transcription of genes kinase domain Interaction inhibit. S, Q, A HIPK1 encoding ribosomal proteins, thus providing a signaling SIM Caspase mechanism connecting environmental stress signaling i S AL to ribosome biogenesis. Another Yak1 substrate protein -Ub involved in the control of ribosomal protein synthesis is kinase domain Interaction S, Q, A HIPK3 SIM Ifh1. While the direct phosphorylation can be recapitu- Caspase lated using in vitro kinase assays (Kim et al., 2011), the AL functional consequences of this phosphorylation remain kinase domain HIPK4 to be studied. A genetic interaction screen revealed the necessity of Yak1 for the mutual binding of the chromatin 100 aa assembly factors Msi1 and Cac1 (Pratt et al., 2007). Further Figure 3 Domain structure of HIPK family members. The various transcriptional regulators found in a proteomic screen domains including the inhibitory (inhibit.) domain are indicated. include the dual function helix-loop-helix protein Cbf1, The positions of the activation loop (AL) and SUMO (S) attachment subunits of the RNA polymerase II mediator complex and sites are displayed. For further details, see text. a subunit of the chromatin-modifying COMPASS (Complex Proteins Associated with Set1)