Cell-cycle control protein expression is disrupted in anogenital condylomata infected with low-risk human papillomavirus types.

OBJECTIVE: To investigate how low-risk human papillomavirus (HPV) infection disrupts cell cycle control. MATERIALS AND METHODS: A series of anogenital condylomata acuminata was analyzed by immunohistochemistry (IHC) for cell cycle protein expression and by polymerase chain reaction and in situ hybridization for HPV type and distribution. RESULTS: Of the 27 condylomata analyzed, 17 contained HPV6 DNA, 8 contained HPV11 DNA and 2 were HPV-negative. Compared with adjacent normal squamous epithelium, there was marked up-regulation of Ki67, cyclin A, and p21 expression in HPV-infected epithelium, particularly in suprabasal keratinocytes. Cyclin E expression mapped to areas with viral DNA amplification. Cyclin D1 expression was largely confined to basal and parabasal cells but was more widespread in the condylomata than in normal epithelium. Only low-level expression of cyclin B was observed. Dual IHC confirmed that expression of p21 was considerably more widespread than p53, consistent with p53-independent expression of p21 in suprabasal keratinocytes in HPV-infected epithelium. Ki67 was expressed throughout the whole thickness of the epithelium in localized areas that were confirmed by dual IHC/in situ hybridization to map to areas of viral DNA amplification. CONCLUSIONS: These data show that cell-cycle disruption as a result of low-risk HPV infection is similar to that reported for productive high-risk HPV infection, suggesting that the life cycles of these 2 viral groups in suprabasal keratinocytes is similar. The reexpression of Ki67 in areas of viral DNA amplification suggests that this protein may play a role in HPV DNA replication.

Use of environmentally enriched housing for rats with spinal cord injury: the need for standardization.

A variety of rehabilitation methods that increase social interaction and locomotor activity are reported to yield positive benefits in humans and animals with spinal cord injury (SCI). Environmental enrichment often incorporates group housing, increased cage size, and objects to increase social interaction and stimulate locomotor activity of animals. Others have reported that adult rats housed in enriched environments immediately after moderate contusion thoracic SCI show improvements in locomotion, but not in neurotransmission through or anatomy at the SCI site. In the present study, in contrast to previous reports, environmental enrichment did not improve the locomotion of rats with contusion thoracic SCI. Furthermore, as in previous reports, improvements were not observed for either electrophysiologic measures of neurotransmission through (transcranial magnetic motor-evoked potentials) and caudal to (magnetic-evoked interlimb reflex) the injury site or the amount of spared white matter at the epicenter. Determining the effectiveness of environmental enrichment to improve locomotor recovery in the SCI model requires standardization of housing procedures, outcome measures, and analyses.

Features in the N and C termini of the MAPK-interacting kinase Mnk1 mediate its nucleocytoplasmic shuttling.

Eukaryotic initiation factor eIF4E binds to the 5'-cap structure of the mRNA and also to the molecular scaffold protein eIF4G. eIF4E is a phosphoprotein, and the kinases that act on it have been identified as the MAPK-interacting kinases Mnk1 and Mnk2. Mnk1/2 also bind to the scaffold protein eIF4G. The N-terminal region of Mnk1 has previously been shown to bind to importin alpha, a component of the nuclear transport machinery, although Mnk1 itself is cytoplasmic. Here we identify a CRM1-type nuclear export motif in the C-terminal part of Mnk1. Substitution of hydrophobic residues in this motif results in Mnk1 becoming nuclear. This has allowed us to study the features of Mnk1 that are involved in its transport to the nucleus. This process requires part, but not all, of a polybasic region near the N terminus of Mnk1. Residues required for nuclear transport are also required for its interaction with importin alpha. This polybasic region also serves a second function in that it is required for the binding of Mnk1 to eIF4G, although the residues involved in this interaction are not identical to those involved in the binding of Mnk1 to importin alpha. Interaction of Mnk1 with eIF4G promotes the phosphorylation of eIF4E. Mutations that reduce the binding of Mnk1 to eIF4G in vivo and in vitro also decrease the ability of Mnk1 to enhance eIF4E phosphorylation in vivo, underlining the importance of the eIF4G-Mnk1 interaction in this process.

Localisation and regulation of the eIF4E-binding protein 4E-BP3.

The cap-binding protein eIF4E-binding protein 3 (4E-BP3) was identified some years ago, but its properties have not been investigated in detail. In this report, we investigated the regulation and localisation of 4E-BP3. We show that 4E-BP3 is present in the nucleus as well as in the cytoplasm in primary T cells, HEK293 cells and HeLa cells. 4E-BP3 was associated with eIF4E in both cell compartments. Furthermore, 4E-BP3/eIF4E association in the cytoplasm was regulated by serum or interleukin-2 starvation in the different cell types. Rapamycin did not affect the association of eIF4E with 4E-BP3 in the cytoplasm or in the nucleus.

A direct inhibitory action of prostaglandins upon ACTH secretion at the late stages of the secretory pathway of AtT-20 cells.

1. The mouse AtT-20/D16-16 anterior pituitary tumour cell line was used as a model system for the study of the effects of prostaglandins upon the late stages of the adrenocorticotrophin (ACTH) secretory pathway. 2. Calcium (1 nM - 100 microM), guanosine-5'-O-(3-thiotriphosphate) (GTP-gamma-S) (1 - 100 microM) and mastoparan (1 and 10 microM) all stimulated ACTH secretion from permeabilized AtT-20 cells in a concentration-dependent manner. GTP-gamma-S and mastoparan stimulated ACTH secretion from permeabilized cells in the absence of calcium. Co-incubation with prostaglandins E(1) and E(2) (PGE(1), PGE(2)) (10 microM) but not prostaglandin F(2 alpha) (PGF(2 alpha)) (10 microM) significantly inhibited calcium-, GTP-gamma-S and mastoparan-evoked secretion by 30 - 50%. 3. The effects of PGE(1) and PGE(2) upon GTP-gamma-S (100 microM)-, calcium (10 microM)- and mastoparan (10 microM)-evoked secretion were concentration-dependent. PGE(1) significantly inhibited GTP-gamma-S- and calcium-evoked secretion at concentrations of PGE(1) above 1 microM but mastoparan-evoked secretion only at the highest concentration of PGE(1) investigated (10 microM). PGE(2) was much more potent than PGE(1) and significantly inhibited GTP-gamma-S- and calcium-evoked secretion at 10 nM and above and mastoparan-evoked secretion above 1 microM. 4. The inhibitory effects of PGE(1) and PGE(2) upon calcium-, GTP-gamma-S- and mastoparan-stimulated ACTH secretion from permeabilized cells were pertussis toxin (PTX) sensitive. 5. In intact cells PGE(1), PGE(2) and PGF(2 alpha) (1 nM - 10 microM) acting singly had little or no effect upon ACTH secretion. However, only PGE(2) (1 nM - 10 microM) significantly inhibited corticotrophin-releasing factor-41 (CRF-41) (100 nM)-evoked secretion in a concentration dependent manner. 6. The present study finds that prostaglandins of the E series exert an inhibitory action, via a pertussis toxin-sensitive GTP-binding (G)-protein, in the late stages of the ACTH secretory pathway distal to the G-exocytosis (Ge)/calcium point of control.