The role of voice input for human-machine communication.

Optimism is growing that the near future will witness rapid growth in human-computer interaction using voice. System prototypes have recently been built that demonstrate speaker-independent real-time speech recognition, and understanding of naturally spoken utterances with vocabularies of 1000 to 2000 words, and larger. Already, computer manufacturers are building speech recognition subsystems into their new product lines. However, before this technology can be broadly useful, a substantial knowledge base is needed about human spoken language and performance during computer-based spoken interaction. This paper reviews application areas in which spoken interaction can play a significant role, assesses potential benefits of spoken interaction with machines, and compares voice with other modalities of human-computer interaction. It also discusses information that will be needed to build a firm empirical foundation for the design of future spoken and multimodal interfaces. Finally, it argues for a more systematic and scientific approach to investigating spoken input and performance with future language technology.

The E5 gene product of rhesus papillomavirus is an activator of endogenous Ras and phosphatidylinositol-3′-kinase in NIH 3T3 cells

We examined the effect of two rhesus papillomavirus 1 (RhPV) oncogenes on cytokine-induced signal transduction pathways leading to the possible activation of Ras protein (p21ras) and phosphatidylinositol kinase. p21ras in both the activated (GTP-bound) and inactivated (GDP-bound) states were quantitated. NIH 3T3 cell lines expressing the RhPV 1 E5 gene or epidermal growth factor receptor cDNA had about a sixfold higher ratio of p21ras-bound GTP to p21ras-bound GDP as compared with parental NIH 3T3 cells or a cell line expressing the RhPV 1 E7 gene under normal culture conditions, yet expressed similar levels of p21ras. Quiescent cells had dramatically reduced levels of activated p21ras, except those containing RhPV 1 E7. Levels were restored by stimulation with epidermal growth factor or platelet-derived growth factor. Both epidermal growth factor and platelet-derived growth factor receptor of RhPV 1 E5- and E7-containing cells responded to cytokine stimulation. Endogenous phosphatidylinositol-3′-kinase was up-regulated in NIH 3T3 cells transformed with the E5 genes of RhPV 1 and bovine papillomavirus 1. These results suggest that E5 genes of papillomaviruses play a major role in the regulation of transduction pathways.

NADPH-oxidase and a hydrogen peroxide-sensitive K+ channel may function as an oxygen sensor complex in airway chemoreceptors and small cell lung carcinoma cell lines

Pulmonary neuroepithelial bodies (NEB) are widely distributed throughout the airway mucosa of human and animal lungs. Based on the observation that NEB cells have a candidate oxygen sensor enzyme complex (NADPH oxidase) and an oxygen-sensitive K+ current, it has been suggested that NEB may function as airway chemoreceptors. Here we report that mRNAs for both the hydrogen peroxide sensitive voltage gated potassium channel subunit (KH2O2) KV3.3a and membrane components of NADPH oxidase (gp91phox and p22phox) are coexpressed in the NEB cells of fetal rabbit and neonatal human lungs. Using a microfluorometry and dihydrorhodamine 123 as a probe to assess H2O2 generation, NEB cells exhibited oxidase activity under basal conditions. The oxidase in NEB cells was significantly stimulated by exposure to phorbol esther (0.1 μM) and inhibited by diphenyliodonium (5 μM). Studies using whole-cell voltage clamp showed that the K+ current of cultured fetal rabbit NEB cells exhibited inactivating properties similar to KV3.3a transcripts expressed in Xenopus oocyte model. Exposure of NEB cells to hydrogen peroxide (H2O2, the dismuted by-product of the oxidase) under normoxia resulted in an increase of the outward K+ current indicating that H2O2 could be the transmitter modulating the O2-sensitive K+ channel. Expressed mRNAs or orresponding protein products for the NADPH oxidase membrane cytochrome b as well as mRNA encoding KV3.3a were identified in small cell lung carcinoma cell lines. The studies presented here provide strong evidence for an oxidase-O2 sensitive potassium channel molecular complex operating as an O2 sensor in NEB cells, which function as chemoreceptors in airways and in NEB related tumors. Such a complex may represent an evolutionary conserved biochemical link for a membrane bound O2-signaling mechanism proposed for other cells and life forms.

The Epstein–Barr virus oncogene product latent membrane protein 1 engages the tumor necrosis factor receptor-associated death domain protein to mediate B lymphocyte growth transformation and activate NF-κB

The Epstein–Barr virus latent membrane protein 1 (LMP1) is essential for the transformation of B lymphocytes into lymphoblastoid cell lines. Previous data are consistent with a model that LMP1 is a constitutively activated receptor that transduces signals for transformation through its carboxyl-terminal cytoplasmic tail. One transformation effector site (TES1), located within the membrane proximal 45 residues of the cytoplasmic tail, constitutively engages tumor necrosis factor receptor-associated factors. Signals from TES1 are sufficient to drive initial proliferation of infected resting B lymphocytes, but most lymphoblastoid cells infected with a virus that does not express the 155 residues beyond TES1 fail to grow as long-term cell lines. We now find that mutating two tyrosines to an isoleucine at the carboxyl end of the cytoplasmic tail cripples the ability of EBV to cause lymphoblastoid cell outgrowth, thereby marking a second transformation effector site, TES2. A yeast two-hybrid screen identified TES2 interacting proteins, including the tumor necrosis factor receptor-associated death domain protein (TRADD). TRADD was the only protein that interacted with wild-type TES2 and not with isoleucine-mutated TES2. TRADD associated with wild-type LMP1 but not with isoleucine-mutated LMP1 in mammalian cells, and TRADD constitutively associated with LMP1 in EBV-transformed cells. In transfection assays, TRADD and TES2 synergistically mediated high-level NF-κB activation. These results indicate that LMP1 appropriates TRADD to enable efficient long-term lymphoblastoid cell outgrowth. High-level NF-κB activation also appears to be a critical component of long-term outgrowth.

CYR61, a product of a growth factor-inducible immediate early gene, promotes angiogenesis and tumor growth

CYR61 is a secreted, cysteine-rich, heparin-binding protein encoded by a growth factor-inducible immediate–early gene. Acting as an extracellular, matrix-associated signaling molecule, CYR61 promotes the adhesion of endothelial cells through interaction with the integrin αVβ3 and augments growth factor-induced DNA synthesis in the same cell type. In this study, we show that purified CYR61 stimulates directed migration of human microvascular endothelial cells in culture through an αVβ3-dependent pathway and induces neovascularization in rat corneas. Both the chemotactic and angiogenic activities of CYR61 can be blocked by specific anti-CYR61 antibodies. Whereas most human tumor-derived cell lines tested express CYR61, the gastric adenocarcinoma cell line RF-1 does not. Expression of the CYR61 cDNA under the regulation of a constitutive promoter in RF-1 cells significantly enhances the tumorigenicity of these cells as measured by growth in immunodeficient mice, resulting in tumors that are larger and more vascularized than those produced by control RF-1 cells. Taken together, these results identify CYR61 as an angiogenic inducer that can promote tumor growth and vascularization; the results also suggest potential roles for CYR61 in physiologic and pathologic neovascularization.

Chromatin-bound PCNA complex formation triggered by DNA damage occurs independent of the ATM gene product in human cells

Proliferating cell nuclear antigen (PCNA), a processivity factor for DNA polymerases δ and ɛ, is involved in DNA replication as well as in diverse DNA repair pathways. In quiescent cells, UV light-induced bulky DNA damage triggers the transition of PCNA from a soluble to an insoluble chromatin-bound form, which is intimately associated with the repair synthesis by polymerases δ and ɛ. In this study, we investigated the efficiency of PCNA complex formation in response to ionizing radiation-induced DNA strand breaks in normal and radiation-sensitive Ataxia telangiectasia (AT) cells by immunofluorescence and western blot techniques. Exposure of normal cells to γ-rays rapidly triggered the formation of PCNA foci in a dose-dependent manner in the nuclei and the PCNA foci (40–45%) co-localized with sites of repair synthesis detected by bromodeoxyuridine labeling. The chromatin-bound PCNA gradually declined with increasing post-irradiation times and almost reached the level of unirradiated cells by 6 h. The PCNA foci formed after γ-irradiation was resistant to high salt extraction and the chromatin association of PCNA was lost after DNase I digestion. Interestingly, two radiosensitive primary fibroblast cell lines, derived from AT patients harboring homozygous mutations in the ATM gene, displayed an efficient PCNA redistribution after γ-irradiation. We also analyzed the PCNA complex induced by a radiomimetic agent, Bleomycin (BLM), which produces predominantly single- and double-strand DNA breaks. The efficiency and the time course of PCNA complex induced by BLM were identical in both normal and AT cells. Our study demonstrates for the first time that the ATM gene product is not required for PCNA complex assembly in response to DNA strand breaks. Additionally, we observed an increased interaction of PCNA with the Ku70 and Ku80 heterodimer after DNA damage, suggestive of a role for PCNA in the non-homologous end-joining repair pathway of DNA strand breaks.

The immunodominant major histocompatibility complex class I-restricted antigen of a murine colon tumor derives from an endogenous retroviral gene product.

Tumors express peptide antigens capable of being recognized by tumor-specific cytotoxic T lymphocytes (CTL). Immunization of mice with a carcinogen-induced colorectal tumor, CT26, engineered to secrete granulocyte/macrophage colony-stimulating factor, routinely generated both short-term and long-term CTL lines that not only lysed the parental tumor in vitro, but also cured mice of established tumor following adoptive transfer in vivo. When either short-term or long-term CTL lines were used to screen peptides isolated from CT26, one reverse-phase high performance liquid chromatography peptide fraction consistently sensitized a surrogate target for specific lysis. The bioactivity remained localized within one fraction following multiple purification procedures, indicating that virtually all of the CT26-specific CTL recognized a single peptide. This result contrasts with other tumor systems, where multiple bioactive peptide fractions have been detected. The bioactive peptide was identified as a nonmutated nonamer derived from the envelope protein (gp70) of an endogenous ecotropic murine leukemia provirus. Adoptive transfer with CTL lines specific for this antigen demonstrated that this epitope represents a potent tumor rejection antigen. The selective expression of this antigen in multiple non-viral-induced tumors provides evidence for a unique class of shared immunodominant tumor associated antigens as targets for antitumor immunity.

The ocular albinism type 1 gene product is a membrane glycoprotein localized to melanosomes.

Ocular albinism type 1 (OA1) is an inherited disorder characterized by severe reduction of visual acuity, photophobia, and retinal hypopigmentation. Ultrastructural examination of skin melanocytes and of the retinal pigment epithelium reveals the presence of macromelanosomes, suggesting a defect in melanosome biogenesis. The gene responsible for OA1 is exclusively expressed in pigment cells and encodes a predicted protein of 404 aa displaying several putative transmembrane domains and sharing no similarities with previously identified molecules. Using polyclonal antibodies we have identified the endogenous OA1 protein in retinal pigment epithelial cells, in normal human melanocytes and in various melanoma cell lines. Two forms of the OA1 protein were identified by Western analysis, a 60-kDa glycoprotein and a doublet of 48 and 45 kDa probably corresponding to unglycosylated precursor polypeptides. Upon subcellular fractionation and phase separation with the nonionic detergent Triton X-114, the OA1 protein segregated into the melanosome-rich fraction and behaved as an authentic integral membrane protein. Immunofluorescence and immunogold analyses on normal human melanocytes confirmed the melanosomal membrane localization of the endogenous OA1 protein, consistent with its possible involvement in melanosome biogenesis. The identification of a novel melanosomal membrane protein involved in a human disease will provide insights into the mechanisms that control the cell-specific pathways of subcellular morphogenesis.