Massive upregulation of the Fas ligand in lpr and gld mice: implications for Fas regulation and the graft-versus-host disease-like wasting syndrome.

Fas-deficient lpr and gld mice develop lymphadenopathy due to the accumulation of T cells with an unusual double negative (DN) (CD4-CD8-) phenotype. Previous studies have shown that these abnormal cells are capable of inducing redirected lysis of certain Fc receptor-positive target cells. Since the Fas ligand (FasL) has recently been shown to be partly responsible for T cell-mediated cytotoxicity, lymph node cells from lpr and gld mice were examined for the expression of FasL mRNA. Northern blot analysis revealed that lymph node cells obtained from lpr and gld mice had a striking increase in the level of expression of FasL mRNA predominantly due to expression in the DN T cells. Furthermore, lpr, but not gld lymph node cells killed the B cell line, A20, in a Fas-dependent manner. These findings indicate that Fas mutations result in a massive up-regulation of FasL which, most likely, results from repetitive exposure to (self) antigen. This phenomenon could explain the lpr-induced wasting syndrome observed when lpr bone marrow-derived cells are adoptively transferred to wild-type recipients.

Conserved helix 7 tyrosine functions as an activation relay in the serotonin 5HT(2C) receptor.

The function of the helix VII Tyr in the conserved Asn-Pro-X-X-Tyr segment of rhodopsin-like G protein coupled receptors has been investigated in many receptors. Various effects of site-directed mutation of this locus have been found, including altered coupling, sequestration and agonist affinity. We report the first constitutively active mutations of this Tyr. In the serotonin 5HT(2C) receptor, substituting Ala or Cys for Tyr resulted in a marked increase in the basal level of inositol phosphate accumulation in transfected COS-1 cells. This constitutive signaling was abolished by the inverse agonist SB206553. Introducing Phe at this locus eliminated both basal and agonist-stimulated signaling. All three mutant receptors showed an increase in binding affinity for the structurally dissimilar agonists 5-hydroxytryptamine (5HT), (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), and quipazine, suggesting that both the activating and inactivating mutations stabilize a high affinity state. These results implicate the conserved Tyr in the conformational rearrangements that occur during agonist complexing and receptor activation.

Pulmonary hemorrhage following intravenous streptokinase for acute myocardial infarction.

A 50-year-old man was given 1.2 million units of intravenous streptokinase 3 hours after the onset of a hyperacute inferior myocardial infarction. He had been treated for pneumonia 4 weeks previously. Five days after thrombolytic therapy, he developed a massive hemoptysis. The implications of this side effect are discussed.

Epstein-Barr nuclear antigen leader protein coactivates transcription through interaction with histone deacetylase 4.

Epstein-Barr nuclear antigen (EBNA) leader protein (EBNALP) coactivates promoters with EBNA2 and is important for Epstein-Barr virus immortalization of B cells. Investigation of the role of histone deacetylases (HDACs) in EBNALP and EBNA2 promoter regulation has now identified EBNALP and EBNA2 to be associated with HDAC4 in a lymphoblastoid cell line. Furthermore, a transcription-deficient EBNALP point mutant did not associate with HDAC4. HDAC4 and 5 overexpression repressed EBNA2 activation and EBNALP coactivation, whereas other HDACs had little effect. Moreover, EBNALP expression decreased nuclear HDAC4. Expression of 14-3-3 anchors HDAC4 in the cytoplasm, increased EBNALP effects, and reversed HDAC4 or 5 repression. HDAC4 reversal depended on the HDAC4 nuclear export sequence. Consistent with EBNALP coactivation being mediated by nuclear HDAC4 depletion, HDAC4 overexpression increased nuclear HDAC4 and specifically repressed EBNA2-dependent activation as well as EBNALP-dependent coactivation. Also, EBNALP, HDAC4, and 14-3-3 could be immunoprecipitated in a single complex. Thus, these data strongly support a model in which EBNALP coactivates transcription by relocalizing HDAC4 and 5 from EBNA2 activated promoters to the cytoplasm. The observed EBNALP effects are likely also in part through HDAC5, which is highly homologous to HDAC4.