Thomas Küntziger

University of Oslo · Department of Oral Biology

Faithful chromosome segregation during mitosis relies on a proofreading mechanism that monitors proper kinetochore-microtubule attachments. The spindle assembly checkpoint (SAC) is based on the concerted action of numerous components that maintain a repressive signal inhibiting transition into anaphase until all chromosomes are attached. Here we sh...

Maintenance of chromatin homeostasis involves proper delivery of histone variants to the genome. The interplay between different chaperones regulating the supply of histone variants to distinct chromatin domains is largely undeciphered. We report here a role of promyelocytic leukemia (PML) protein in routing histone variant H3.3 to chromatin and in...

Replication-independent chromatin deposition of histone variant H3.3 is mediated by several chaperones. We report a multi-step targeting of newly synthesized epitope-tagged H3.3 to chromatin via PML bodies. H3.3 is recruited to PML bodies in a DAXX--dependent manner, a process facilitated by ASF1A. DAXX is required for enrichment of ATRX, but not A...

Adrenergic stimulation of adipocytes yields a cAMP signal that activates protein kinase A (PKA). PKA phosphorylates perilipin, a protein localized on the surface of lipid droplets that serves as a gatekeeper to regulate access of lipases converting stored triglycerides to free fatty acids and glycerol in a phosphorylation-dependent manner. Here, we...

The function of protein phosphatase 1 nuclear-targeting subunit (PNUTS)--one of the most abundant nuclear-targeting subunits of protein phosphatase 1 (PP1c)--remains largely uncharacterized. We show that PNUTS depletion by small interfering RNA activates a G2 checkpoint in unperturbed cells and prolongs G2 checkpoint and Chk1 activation after ioniz...

Cyclic di-adenosine monophosphate (c-di-AMP) has emerged as an important bacterial signaling molecule that functions both as an intracellular second messenger in bacterial cells and an extracellular ligand involved in bacteria-host cross-talk. In this study, we identify and characterize proteins involved in controlling the c-di-AMP concentration in...

Optic atrophy 1 (OPA1) is the A-kinase anchoring protein (AKAP) targeting the pool of PKA responsible for perilipin 1 phosphorylation, a gatekeeper for lipolysis. However, the involvement of OPA1-bound PKA in the downstream regulation of lipolysis is unknown. Here, we show upregulation and relocation of OPA1 from mitochondria to lipid droplets duri...

Cover: The cover image, by Eva Bártová et al., is based on the Article Localized Movement and Levels of 53BP1 Protein Are Changed by γ-irradiation in PML Deficient Cells, DOI: 10.1002/jcb.25551.

We studied epigenetics, distribution pattern, kinetics and diffusion of proteins recruited to spontaneous and γ-radiation-induced DNA lesions. We showed that PML deficiency leads to an increased number of DNA lesions, which was accompanied by changes in histone signature. In PML wt cells, we observed two mobile fractions of 53BP1 protein with disti...

Histone variant H3.3 is deposited in chromatin at active sites, telomeres and pericentric heterochromatin by distinct chaperone, but the mechanisms of regulation and coordination of chaperone-mediated H3.3 loading remain largely unknown. We show here that the chromatin-associated oncoprotein DEK regulates differential HIRA- and DAAX/ATRX-dependent...

Background The aquatic birnavirus infectious pancreatic necrosis virus (IPNV) causes infectious pancreatic necrosis (IPN), a severe disease in farmed salmonid fish. IPNV has a very broad host range and infects many different species of fish as well as molluscs and crustaceans. Investigation of the host reservoir of a virus may reveal important mole...

In response to DNA damaging agents and endogenous DNA lesions, human cells activate signaling cascades and repair mechanisms to help maintain genomic integrity. Phosphorylation plays a major role in DNA damage signaling, and the role of Ser/Thr kinases, including ATM, ATR, CHK1, CHK2 and DNA-PK, is particularly well documented. While these kinases...

In contrast to canonical histones, histone variant H3.3 is incorporated into chromatin in a replication-independent manner. Posttranslational modifications of H3.3 have been identified; however, the epigenetic environment of incorporated H3.3 is unclear. We have investigated the genomic distribution of epitope-tagged H3.3 in relation to histone mod...

A-kinase-anchoring protein 149 (AKAP149) is a membrane protein of the mitochondrial and endoplasmic reticulum/nuclear envelope network. AKAP149 controls the subcellular localization and temporal order of protein phosphorylation by tethering protein kinases and phosphatases to these compartments. AKAP149 also includes an RNA-binding K homology (KH)...

A-Kinase anchoring proteins (AKAPs) control the subcellular localization and temporal specificity of protein phosphorylation mediated by cAMP-dependent protein kinase. AKAP149 (AKAP1) is found in mitochondria and in the endoplasmic reticulum-nuclear envelope network where it anchors protein kinases, phosphatases, and a phosphodiesterase. AKAP149 ha...

This chapter examines procedures for reprogramming somatic nuclei and cells with cell extracts. About 293T human fibroblasts are cultured on glass coverslips. Resuspend the cells in 10 ml ice-cold cell lysis buffer (CLB). It is preferable to use a graduated 15-ml conical tube to estimate the cell volume after sedimentation. Once lysis is achieved i...

Reformation of the nuclear envelope at the end of mitosis involves the recruitment of the B-type lamin phosphatase PP1 to nuclear membranes by A-kinase anchoring protein AKAP149. PP1 remains associated to AKAP149 throughout G1 but dissociates from AKAP149 when AKAP149 is phosphorylated at the G1/S transition. We examine here the role of phosphoryla...

Directly turning a somatic cell type into another would be beneficial for producing replacement cells for therapeutic purposes. To this end, novel cell reprogramming strategies are being developed. We describe here methods for functionally reprogramming a somatic cell using an extract derived from another somatic cell type. The procedure involves r...

We demonstrate a cell extract-based, genome-wide and heritable reprogramming of gene expression in vitro. Kidney epithelial 293T cells have previously been shown to take on T cell properties following a brief treatment with an extract of Jurkat T cells. We show here that 293T cells exposed for 1 h to a Jurkat cell extract undergo genome-wide, targe...

PP1 (protein phosphatase-1) is a serine/threonine phosphatase involved in mitosis exit and chromosome decondensation. In the present study, we characterize the subcellular and subnuclear localization of PNUTS (PP1 nuclear targeting subunit), a nuclear regulatory subunit of PP1, and report a stimulatory role of PNUTS in the decondensation of prometa...

This chapter addresses the transdifferentiation potential of somatic cells and illustrates the functional reprogramming of somatic cells using extracts derived from another somatic cell type. Novel ways of redirecting the fate of differentiated somatic cells, a process known as transdifferentiation, are actively being investigated. Methods for tran...

The specificity of action of protein kinases and phosphatases can be achieved by their recruitment into multiprotein complexes at discrete subcellular loci. One class of molecules targeting kinases and phosphatases within single complexes includes A-kinase (PKA) anchoring proteins or AKAPs. Interestingly, AKAPs not only anchor enzymes but may also...

We have developed a somatic cell-free system that remodels chromatin and activates gene expression in heterologous differentiated nuclei. Extracts of stimulated human T cells elicit chromatin binding of transcriptional activators of the interleukin-2 (IL-2) gene, anchoring and activity of a chromatin-remodeling complex and hyperacetylation of the I...

Stathmin/Op18 destabilizes microtubules in vitro and regulates microtubule polymerization in vivo. Both a microtubule catastrophe-promoting activity and a tubulin sequestering activity were demonstrated for stathmin in vitro, and both could contribute to microtubule depolymerization in vivo. Stathmin activity can be turned down by extensive phospho...

Stathmin/Op 18 is a microtubule (MT) dynamics-regulating protein that has been shown to have both catastrophe-promoting and tubulin-sequestering activities. The level of stathmin/Op18 phosphorylation was proved both in vitro and in vivo to be important in modulating its MT-destabilizing activity. To understand the in vivo regulation of stathmin/Op1...

Centrosome reproduction by duplication is essential for the bipolarity of cell division, but the molecular basis of this process is still unknown. Mutations in Saccharomyces cerevisiae CDC31 gene prevent the duplication of the spindle pole body (SPB). The product of this gene belongs to the calmodulin super-family and is concentrated at the half br...

This chapter discusses the functional description of the centrosome as well as two main aspects of centrosome reproduction and the parthenogenetic activity of the centrosome: (1) the molecular mechanisms underlying centrosome biogenesis itself and (2) the coordination between the nuclear cycle and the centrosome cycle. Isolating centrosomes reveals...