microRNA-10b enhances pancreatic cancer cell invasion by suppressing TIP30 expression and promoting EGF and TGF-ß actions.

This corrects the article DOI: 10.1038/onc.2013.405.

microRNA-10b enhances pancreatic cancer cell invasion by suppressing TIP30 expression and promoting EGF and TGF-ß actions.

Increased microRNA-10b (miR-10b) expression in the cancer cells in pancreatic ductal adenocarcinoma (PDAC) is a marker of disease aggressiveness. In the present study, we determined that plasma miR-10b levels are significantly increased in PDAC patients by comparison with normal controls. By gene profiling, we identified potential targets downregulated by miR-10b, including Tat-interacting protein 30 (TIP30). Immunoblotting and luciferase reporter assays confirmed that TIP30 was a direct miR-10b target. Downregulation of TIP30 by miR-10b or siRNA-mediated silencing of TIP30 enhanced epidermal growth factor (EGF)-dependent invasion. The actions of miR-10b were abrogated by expressing a modified TIP30 cDNA resistant to miR-10b. EGF-induced EGF receptor (EGFR) tyrosine phosphorylation and extracellular signal-regulated kinase phosphorylation were enhanced by miR-10b, and these effects were mimicked by TIP30 silencing. The actions of EGF in the presence of miR-10b were blocked by EGFR kinase inhibition with erlotinib and by dual inhibition of PI3K (phosphatidylinositol 3'-kinase) and MEK. Moreover, miR-10b, EGF and transforming growth factor-beta (TGF-ß) combined to markedly increase cell invasion, and this effect was blocked by the combination of erlotinib and SB505124, a type I TGF-ß receptor inhibitor. miR-10b also enhanced the stimulatory effects of EGF and TGF-ß on cell migration and epithelial-mesenchymal transition (EMT) and decreased the expression of RAP2A, EPHB2, KLF4 and NF1. Moreover, miR-10b overexpression accelerated pancreatic cancer cell (PCC) proliferation and tumor growth in an orthotopic model. Thus, plasma miR-10b levels may serve as a diagnostic marker in PDAC, whereas intra-tumoral miR-10b promotes PCC proliferation and invasion by suppressing TIP30, which enhances EGFR signaling, facilitates EGF-TGF-ß cross-talk and enhances the expression of EMT-promoting genes, whereas decreasing the expression of several metastasis-suppressing genes. Therefore, therapeutic targeting of miR-10b in PDAC may interrupt growth-promoting deleterious EGF-TGF-ß interactions and antagonize the metastatic process at various levels.

MHC I-dependent antigen presentation is inhibited by poliovirus protein 3A.

The effects of poliovirus 3A protein expression and poliovirus infection on the presentation of hepatitis C virus antigens in cultured chimpanzee cells were examined. Expression of poliovirus 3A protein inhibits protein secretion when expressed in isolation and was sufficient to protect chimpanzee cells from lysis by hepatitis C virus-specific cytotoxic T cells in standard (51)Cr-release assays. Poliovirus infection also inhibited antigen presentation, as determined by decreased cytotoxic T cell activation. A mutation in 3A that abrogates the inhibition of protein secretion also abolished the effects of poliovirus on antigen presentation. These results demonstrate that the inhibition of secretion observed in poliovirus-infected cells substantially reduces the presentation of new antigens on the cell surface. These observations may reflect a general mechanism by which nonenveloped viruses such as poliovirus and other viruses that do not require a functional protein secretory apparatus can evade detection by the cellular immune response.

Morphogenesis and dynamics of the yeast Golgi apparatus.

A kinetic and morphometric study was conducted with the electron microscope to clarify the biogenesis and structural diversity of the Golgi apparatus in the yeast Saccharomyces cerevisiae. Secretion was synchronized by inhibiting protein synthesis and/or by subjecting thermosensitive secretory mutants to double temperature shifts. Five membrane-bounded structures disappeared or reappeared in an orderly manner at approximately the rate of secretory protein flow. 1) The first detectable post-ER intermediates were very short-lived clusters of small vesicles that appeared next to the endoplasmic reticulum (ER). 2) Their constituent small vesicles were rapidly bridged by membrane tubules in a SEC18-dependent manner, giving short-lived tubular clusters of small vesicles, analogous to mammalian vesicular-tubular clusters. 3) Fine and 4) large nodular networks (coated with the Golgi protein Sec7), and 5) secretory granules. Upon relieving a secretory block, each structure successively reappeared, seemingly by transformation of the previous one. When no secretory cargo was to be transported, these structures were not renewed. They disappeared more than five times faster than some Golgi enzymes such as Och1p, implying that the latter are recycled and perhaps partially retained. Retention could arise from intra-compartmental flow of cargo/carrier, hinted at by the varying calibers within a single nodular network.

Sec7p directs the transitions required for yeast Golgi biogenesis.

Endoplasmic reticulum (ER)-to-Golgi traffic in yeast proceeds by the maturation of membrane compartments from post-ER vesicles to intermediate small vesicle tubular clusters (VTCs) to Golgi nodular membrane networks (Morin-Ganet et al., Traffic 2000; 1: 56-68). The balance between ER and Golgi compartments is maintained by COPII- and COPI-mediated anterograde and retrograde traffic, which are dependent on Sec7p and ARF function. The sec7-4 temperature-sensitive allele is a mutation in the highly conserved Sec7 domain (Sec7d) found in all ARF-guanine nucleotide exchange factor proteins. Post-ER trafficking is rapidly inactivated in sec7-4 mutant yeast at the restrictive temperature. This conditional defect prevented the normal production of VTCs and instead generated Golgi-like tubes emanating from the ER exit sites. These tubes progressively developed into stacked cisternae defining the landmark sec7 mutant phenotype. Consistent with the in vivo results, a Sec7d peptide inhibited ER-to-Golgi transport and displaced Sec7p from its membrane anchor in vitro. The similarities in the consequences of inactivating Sec7p or ARFs in vivo was revealed by genetic disruption of yeast ARFs or by addition of brefeldin A (BFA) to whole cells. These treatments, as in sec7-4 yeast, affected the morphology of membrane compartments in the ER-Golgi transition. Further evidence for Sec7p involvement in the transition for Golgi biogenesis was revealed by in vitro binding between distinct domains of Sec7p with ARFs, COPI and COPII coat proteins. These results suggest that Sec7p coordinates membrane transitions in Golgi biogenesis by directing and scaffolding the binding and disassembly of coat protein complexes to membranes, both at the VTC transition from ER exit sites to form Golgi elements and for later events in Golgi maturation.

An N-end rule destabilization mutant reveals pre-Golgi requirements for Sec7p in yeast membrane traffic.

Sec7 protein (Sec7p) is required for membrane traffic in the yeast secretory pathway. Because Sec7p regulates more than one stage in the pathway, it has been difficult to assign the most proximal requirement for Sec7p action. We have engineered a novel mutant whose Sec7p levels are regulated by growth conditions and by selective protein destabilization according to the N-end rule. Sec7p depletion causes cell growth arrest and accumulation of transport proteins with post-translational modifications indicative of Sec7p dependence for ER-to-Golgi traffic, in addition to the already characterized Golgi requirements. Immuno-EM of sec7 revealed exaggeration of ER and Golgi membranes with protein accumulation in these exaggerated structures, suggesting that these regions may represent staging areas for cargo sorting and vesicle assembly.

Human ARF4 expression rescues sec7 mutant yeast cells.

Vesicle-mediated traffic between compartments of the yeast secretory pathway involves recruitment of multiple cytosolic proteins for budding, targeting, and membrane fusion events. The SEC7 gene product (Sec7p) is a constituent of coat structures on transport vesicles en route to the Golgi complex in the yeast Saccharomyces cerevisiae. To identify mammalian homologs of Sec7p and its interacting proteins, we used a genetic selection strategy in which a human HepG2 cDNA library was transformed into conditional-lethal yeast sec7 mutants. We isolated several clones capable of rescuing sec7 mutant growth at the restrictive temperature. The cDNA encoding the most effective suppressor was identified as human ADP ribosylation factor 4 (hARF4), a member of the GTPase family proposed to regulate recruitment of vesicle coat proteins in mammalian cells. Having identified a Sec7p-interacting protein rather than the mammalian Sec7p homolog, we provide evidence that hARF4 suppressed the sec7 mutation by restoring secretory pathway function. Shifting sec7 strains to the restrictive temperature results in the disappearance of the mutant Sec7p cytosolic pool without apparent changes in the membrane-associated fraction. The introduction of hARF4 to the cells maintained the balance between cytosolic and membrane-associated Sec7p pools. These results suggest a requirement for Sec7p cycling on and off of the membranes for cell growth and vesicular traffic. In addition, overexpression of the yeast GTPase-encoding genes ARF1 and ARF2, but not that of YPT1, suppressed the sec7 mutant growth phenotype in an allele-specific manner. This allele specificity indicates that individual ARFs are recruited to perform two different Sec7p-related functions in vesicle coat dynamics.

Diagnosis and treatment of pneumonia in the nursing home.

Risk factors, diagnosis, ethical considerations, and treatment of pneumonia in a nursing home setting are summarized. Risk factors for pneumonia include age-associated changes, co-morbid conditions, declining general health, and iatrogenic factors. Diagnosis can be challenging in geriatric residents because of atypical presentations and complex underlying diseases. Key features of presentation include rhonchi and confusion. An increased respiratory rate can be a sensitive indicator for pneumonia. Ethical dilemmas include identifying a resident's wishes for treatment in the event of an acute illness and the decision on whether to relocate the resident to an acute care facility. Decisions are based on available resources, medical stability of the resident, unimpeded access to a hospital, and the resident's well-informed decision. The common etiologic pathogen(s) of nursing-home-acquired pneumonia reflect a mixture between community-acquired and hospital-acquired pathogens. Multiple pharmacologic interventions are available, as well as supportive measures including oxygen therapy and hydration. Preventive measures are utilized to decrease the incidence of further infection.

Feedback effects on sequential ordering in humans.

Under various feedback conditions, 38 college undergraduates were asked to rearrange abstract graphic characters on a computer screen, placing them in arbitrarily designated "correct" sequences. Two sets of seven horizontally arrayed stimuli were used. In Experiment 1, subjects in Group 1 learned to arrange the first set under Selection Feedback in which a "+" appeared above each character after it was selected in the correct order and to arrange the second set under Order Feedback in which a correct response produced a copy of the character in its correct ordinal position at the top of the screen. For Group 2 the order of these conditions was reversed. In Experiment 2, for subjects in Group 3, correct responses produced neither of these types of feedback. Subjects in Group 4 received Order Feedback only until the first set was correctly ordered once. Order Feedback was more effective than Selection Feedback during initial acquisition of the first set but not during maintenance; no differences were found for the second set. Only 2 of 9 subjects successfully put the characters in correct sequential order under the No Feedback condition. When, in Experiment 2, Order Feedback was eliminated after the first correctly arranged sequence, the steady-state criteria were met more slowly than in Experiment 1.

Comparing the effects of two correction procedures on human acquisition of sequential behavior patterns.

Thirty-one college undergraduates learned to touch abstract stimuli on a computer screen in arbitrarily designated "correct" sequential orders. Four sets of seven stimuli were used; the stimuli were arrayed horizontally on the screen in random sequences. A correct response (i.e., touching first the stimulus designated as first) resulted in that stimulus appearing near the top of the screen in its correct sequential position (left to right), and remaining there until the end of the trial. Incorrect responses (i.e., touching a stimulus out of sequence) terminated the trial. New trials displayed either the same sequence as the one on which an error had occurred (same-order correction procedure), or a new random sequence (new-order correction procedure). Whenever all responses occurred in the correct sequence, the next trial displayed a new random sequence. Each phase ended when five consecutive correct response sequences occurred. Initially, the same-order correction procedure increased control by the position as well as by the shape of the stimuli; also, it produced more errors, more total trials, more trials to mastery, and more individual patterns of reacquisition than were produced by the new-order procedure.

Understanding cognitive language: The mental idioms in children's talk.

Considerable debate has occurred among behavior analysts about the value of cognitive language for labels or descriptions of phenomena in the analysis of behavior. That value is difficult to assess, however, until a clearer understanding of the definitions of those terms is obtained. To begin that process, this article demonstrates through a series of examples what children mean when they use typical cognitive expressions. One conclusion possible from the results of such an analysis is that cognitive terms describe nothing more than behavior in context, a very behavioral idea. Cognitive expressions may be more suitable to a behavioral analysis than to one derived from the current computer metaphor of cognitive science. The usefulness of these more accurately defined cognitive expressions for the scientific language of behavior analysis is discussed.

Measurement of empathy toward rape victims and rapists.

The purpose of the present study was the construction of the Rape Empathy Scale (RES), designed to measure subjects' empathy toward the rape victim and the rapist in a heterosexual rape situation. The results of psychometric analyses of reliability for both a student and juror sample are presented, in addition to evidence of cross-validation on separate student and juror samples. Significant differences between male and female subjects' RES scores were found, as well as differences between scores of women who had experienced a rape situation (rape victims and rape resisters) and women with no previous exposure to rape. RES scores were predictive of both students' and jurors' ratings of defendant guilt, as well as their recommended sentences for the defendant and their attributions of responsibility for the crime. Furthermore, subjects' RES scores were predictive of their social perceptions of the rape victim and defendant, and male jurors' RES scores were negatively correlated with their reported desire to rape a woman. The results are discussed in relation to the low conviction rate for sexual assault cases and the importance of juror selection as a vehicle for increasing the number of just convictions in rape cases.

Defining applied behavior analysis: an historical analogy.

This article examines two criteria for a definition of applied behavior analysis. The criteria are derived from a 19th century attempt to establish medicine as a scientific field. The first criterion, experimental determinism, specifies the methodological boundaries of an experimental science. The second criterion, philosophic doubt, clarifies the tentative nature of facts and theories derived from those facts. Practices which will advance the science of behavior are commented upon within each criteria. To conclude, the problems of a 19th century form of empiricism in medicine are related to current practices in applied behavior analysis.