Biomedical subjects

Michelle Hughes

Publications and source records attributed to Michelle Hughes.

4 recordsLinked to original sources

PubMed2003

Fas-associated death domain protein and caspase-8 are not recruited to the tumor necrosis factor receptor 1 signaling complex during tumor necrosis factor-induced apoptosis.

Death receptors are a subfamily of the tumor necrosis factor (TNF) receptor subfamily. They are characterized by a death domain (DD) motif within their intracellular domain, which is required for the induction of apoptosis. Fas-associated death domain protein (FADD) is reported to be the universal adaptor used by death receptors to recruit and activate the initiator caspase-8. CD95, TNF-related apoptosis-inducing ligand (TRAIL-R1), and TRAIL-R2 bind FADD directly, whereas recruitment to TNF-R1 is indirect through another adaptor TNF receptor-associated death domain protein (TRADD). TRADD also binds two other adaptors receptor-interacting protein (RIP) and TNF-receptor-associated factor 2 (TRAF2), which are required for TNF-induced NF-kappaB and c-Jun N-terminal kinase activation, respectively. Analysis of the native TNF signaling complex revealed the recruitment of RIP, TRADD, and TRAF2 but not FADD or caspase-8. TNF failed to induce apoptosis in FADD- and caspase-8-deficient Jurkat cells, indicating that these apoptotic mediators were required for TNF-induced apoptosis. In an in vitro binding assay, the intracellular domain of TNF-R1 bound TRADD, RIP, and TRAF2 but did not bind FADD or caspase-8. Under the same conditions, the intracellular domain of both CD95 and TRAIL-R2 bound both FADD and caspase-8. Taken together these results suggest that apoptosis signaling by TNF is distinct from that induced by CD95 and TRAIL. Although caspase-8 and FADD are obligatory for TNF-mediated apoptosis, they are not recruited to a TNF-induced membrane-bound receptor signaling complex as occurs during CD95 or TRAIL signaling, but instead must be activated elsewhere within the cell.

Adaptor Proteins, Signal Transducing↗

PubMed2003

Herpes simplex virus type 1 portal protein UL6 interacts with the putative terminase subunits UL15 and UL28.

The herpes simplex virus type 1 (HSV-1) UL6, UL15, and UL28 proteins are essential for cleavage of replicated concatemeric viral DNA into unit length genomes and their packaging into a preformed icosahedral capsid known as the procapsid. The capsid-associated UL6 DNA-packaging protein is located at a single vertex and is thought to form the portal through which the genome enters the procapsid. The UL15 protein interacts with the UL28 protein, and both are strong candidates for subunits of the viral terminase, a key component of the molecular motor that drives the DNA into the capsid. To investigate the association of the UL6 protein with the UL15 and UL28 proteins, the three proteins were produced in large amounts in insect cells with the baculovirus expression system. Interactions between UL6 and UL28 and between UL6 and UL15 were identified by an immunoprecipitation assay. These results were confirmed by transiently expressing wild-type and mutant proteins in mammalian cells and monitoring their distribution by immunofluorescence. In cells expressing the single proteins, UL6 and UL15 were concentrated in the nuclei whereas UL28 was found in the cytoplasm. When the UL6 and UL28 proteins were coexpressed, UL28 was redistributed to the nuclei, where it colocalized with UL6. In cells producing either of two cytoplasmic UL6 mutant proteins and a functional epitope-tagged form of UL15, the UL15 protein was concentrated with the mutant UL6 protein in the cytoplasm. These observed interactions of UL6 with UL15 and UL28 are likely to be of major importance in establishing a functional DNA-packaging complex at the portal vertex of the HSV-1 capsid.

Animals↗

PubMed2002

Binaural cochlear implants placed during the same operation.

OBJECTIVE: To evaluate the binaural listening advantages for speech in quiet and in noise and to localize sound when independently programmed binaural cochlear implants are used, and to determine whether ears with different hearing ability and duration of profound deafness perform differently with cochlear implants as well as to what extent preimplant psychophysical and physiologic assessment could be predictive of performance. STUDY DESIGN: Prospective study in which patients were prospectively selected to undergo bilateral implantation during a single surgical procedure at a tertiary referral center. All testing was performed with patients using their right, left, or both cochlear implants. Preimplant and intraoperative measures used electrical stimulation at the round window and stimulation through the cochlear implant. RESULTS: Bilateral implantation during the same operation did not cause any postoperative problems such as severe vertigo or ataxia. At 1 year, results of speech testing in quiet demonstrated a binaural advantage for 2 of 10 subjects. Speech-in-noise testing demonstrated that two implants were beneficial for two individuals. All subjects benefited from a head shadow effect when an ear with a better signal-to-noise ratio was available. The ability to localize sound was improved with binaural implants in all subjects. Preimplant psychophysical or physiologic measures were not predictive of eventual speech perception performance. CONCLUSION: Binaural cochlear implants can assist in the localization of sounds and have the potential in some individuals to improve speech understanding in quiet and in noise.

Adult↗

PubMed2000

Interaction of the herpes simplex virus type 1 packaging protein UL15 with full-length and deleted forms of the UL28 protein.

The UL15 and UL28 proteins of herpes simplex virus type 1 are both required for the packaging of replicated viral DNA into the viral capsid. We have expressed UL28 and a functional epitope-tagged form of UL15 in mammalian and insect cells. Immunoprecipitation experiments confirmed that the two proteins can interact. In agreement with previous results, UL15, when expressed alone, entered the nucleus but UL28 remained cytoplasmic. When co-expressed the two proteins co-localized in the nucleus. Six UL28 deletion mutants were constructed and similarly analysed. The results obtained by immunoprecipitation and immunofluorescence were consistent and demonstrate that at least two separate regions of the UL28 polypeptide chain have the ability to interact with UL15. Surprisingly, three of the mutants prevented the UL15 protein from localizing to the cell nucleus, and these were not functional in a transient DNA packaging assay. Of the three UL28 mutant proteins that entered the nucleus with UL15, one containing an internal deletion of 13 amino acids was able to complement a UL28 null mutant in both DNA packaging and virus yield assays, demonstrating that this region of the protein is not essential for function. In addition to interacting with the UL28 protein we also demonstrated that UL15 molecules can interact with each other, and that sequences within the second exon contribute to this interaction.

Active Transport, Cell Nucleus↗