Biomedical subjects

Konrad Krzewski

Publications and source records attributed to Konrad Krzewski.

4 recordsLinked to original sources

PubMed2006

CD28-stimulated ERK2 phosphorylation is required for polarization of the microtubule organizing center and granules in YTS NK cells.

Activation of natural killer (NK) cell cytotoxicity requires adhesion and formation of a conjugate with a susceptible target cell, followed by actin polymerization, and polarization of the microtubule organizing center (MTOC) and cytolytic granules to the NK cell immune synapse. Here, by using the YTS NK cell line as a model, CD28 is shown to be an activating receptor. It signals cytotoxicity in a process dependent on phosphoinositide-3 kinase activation, leading to sustained extracellular signal-regulated kinase 2 (ERK2) phosphorylation. ERK and phospho-ERK localize to microtubule filaments. Neither conjugation with targets nor actin polymerization is affected by blocking ERK2 activation. However, both polarization of the MTOC and cytolytic granules to the synaptic region and NK cell cytotoxicity are strongly reduced by blocking ERK2 activation. A role for the CD28/CD80 interaction in cytotoxicity of human peripheral NK cells also was established. By contrast, lymphocyte function-associated antigen 1 (LFA-1) ligation transduces only a transient ERK2 activation and fails to induce killing in YTS cells. Thus, in YTS cells, a CD28 signal is used to polarize the MTOC and cytolytic granules to the NK cell immune synapse by stimulating sustained ERK2 activation.

Apoptosis↗

PubMed2006

Formation of a WIP-, WASp-, actin-, and myosin IIA-containing multiprotein complex in activated NK cells and its alteration by KIR inhibitory signaling.

The tumor natural killer (NK) cell line YTS was used to examine the cytoskeletal rearrangements required for cytolysis. A multiprotein complex weighing approximately 1.3 mD and consisting of WASp-interacting protein (WIP), Wiskott-Aldrich syndrome protein (WASp), actin, and myosin IIA that formed during NK cell activation was identified. After induction of an inhibitory signal, the recruitment of actin and myosin IIA to a constitutive WIP-WASp complex was greatly decreased. Both actin and myosin IIA were recruited to WIP in the absence of WASp. This recruitment correlated with increased WIP phosphorylation, which was mediated by PKCtheta. Furthermore, the disruption of WIP expression by WIP RNA interference prevented the formation of this protein complex and led to almost complete inhibition of cytotoxic activity. Thus, the multiprotein complex is important for NK cell function, killer cell immunoglobulin-like receptor inhibitory signaling affects proteins involved in cytoskeletal rearrangements, and WIP plays a central role in the formation of the complex and in the regulation of NK cell activity.

Actins↗

PubMed2004

Human nuclear factor kappa B essential modulator mutation can result in immunodeficiency without ectodermal dysplasia.

BACKGROUND: Many receptors rely on the appropriate activation of nuclear factor (NF) kappa B to induce cellular function. This process depends critically on the phosphorylation of the inhibitor of NF-kappa B (I kappa B) by the I kappa B kinase. This targets I kappa B for ubiquitination and degradation, allowing NF-kappa B to translocate to the nucleus, where it can direct transcription. Hypomorphic human mutations affecting one I kappa B kinase component, the NF-kappa B essential modulator (NEMO), result in impaired signaling from receptors required for ectodermal development and immune function. Male subjects with these mutant NEMO molecules have an X-linked syndrome known as ectodermal dysplasia with immunodeficiency, which is characterized by severe infections, with herpesviruses, bacteria, and mycobacterial susceptibility. OBJECTIVE: We sought to genetically and biochemically characterize a patient with a mutant NEMO molecule without ectodermal abnormalities. METHODS: We evaluated NEMO in a patient who had immunodeficiency and atypical mycobacterial infection but normal ectoderm. RESULTS: We identified a novel NEMO mutant causing immunodeficiency without ectodermal dysplasia. The mutation, which altered the exon 9 splice site, was present in cells of ectodermal and hematopoetic origin and resulted in a heterogeneous mixture of mutant and wild-type cDNA species. Immunologic function was variably impaired, with reduced CD40-induced B-cell proliferation, partially reduced NF-kappa B p65 nuclear translocation, and variable Toll-like receptor-induced TNF production. This variability might be explained by an inconsistent ratio of mutant to wild-type NEMO. The lack of any ectodermal phenotype, however, suggested a separation in the hematopoetic and ectodermal function of NEMO that leads to NF-kappa B activation. CONCLUSION: Mutation of the gene encoding NEMO can result in immunodeficiency without ectodermal dysplasia.

Adolescent↗

PubMed2003

Characterization of the anti-DnaJ monoclonal antibodies and their use to compare immunological properties of DnaJ and its human homologue HDJ-1.

Escherichia coli DnaJ (Hsp40) is suspected to participate in rheumatoid arthritis (RA) pathogenesis in humans by an autoimmune process. In this work a set of 6 anti-DnaJ monoclonal antibodies (mAbs) was raised and localization of the epitopes recognized by the mAbs was investigated. Western blotting and enzyme-linked immunosorbent assay (ELISA) experiments showed that the mAbs efficiently bound only native antigen. Using DnaJ mutant proteins with deletions of specified domains and ELISA, we found that AC11 mAb reacted with the best conserved in evolution N-terminal J domain, whereas BB3, EE11, CC5, CC8, and DC7 bound to the C-terminal part after residue 200. Mapping performed with the use of a random peptide library displayed by filamentous phage indicated that (1) AC11 mAb bound to a region between residues 33-48, including D-34 which belongs to the HPD triad, present in all DnaJ homologues, (2) BB3 recognized residues localized in the 204-224 region, (3) EE11 recognized the 291-309 region, (4) CC5--the region 326-359, and (5) CC8--the 346-366 region. All these mAbs, as well as the polyclonal antibodies against the N- or C-terminal domain, bound efficiently to HDJ-1, human Hsp40. These results show the presence of a significant immunological similarity between bacterial DnaJ and human HDJ-1, which is not restricted to the evolutionarily conserved parts of the proteins, and suggest that HDJ-1 could be a possible target of immune response triggered by DnaJ.

Animals↗