Placental Mammals Acquired Functional Sequences in NRK for Regulating the CK2-PTEN-AKT Pathway and Placental Cell Proliferation.

The molecular evolution processes underlying the acquisition of the placenta in eutherian ancestors are not fully understood. Mouse NCK-interacting kinase (NIK)-related kinase (NRK) is expressed highly in the placenta and plays a role in preventing placental hyperplasia. Here, we show the molecular evolution of NRK, which confers its function for inhibiting placental cell proliferation. Comparative genome analysis identified NRK orthologs across vertebrates, which share the kinase and citron homology (CNH) domains. Evolutionary analysis revealed that NRK underwent extensive amino acid substitutions in the ancestor of placental mammals and has been since conserved. Biochemical analysis of mouse NRK revealed that the CNH domain binds to phospholipids, and a region in NRK binds to and inhibits casein kinase-2 (CK2), which we named the CK2-inhibitory region (CIR). Cell culture experiments suggest the following: 1) Mouse NRK is localized at the plasma membrane via the CNH domain, where the CIR inhibits CK2. 2) This mitigates CK2-dependent phosphorylation and inhibition of PTEN and 3) leads to the inhibition of AKT signaling and cell proliferation. Nrk deficiency increased phosphorylation levels of PTEN and AKT in mouse placenta, supporting our hypothesis. Unlike mouse NRK, chicken NRK did not bind to phospholipids and CK2, decrease phosphorylation of AKT, or inhibit cell proliferation. Both the CNH domain and CIR have evolved under purifying selection in placental mammals. Taken together, our study suggests that placental mammals acquired the phospholipid-binding CNH domain and CIR in NRK for regulating the CK2-PTEN-AKT pathway and placental cell proliferation.

Specific interaction of the envelope glycoproteins E1 and E2 with liver heparan sulfate involved in the tissue tropismatic infection by hepatitis C virus.

The first step in the process of infections by the hepatitis C virus (HCV) is attachment to the host cell, which is assumed to be mediated by interaction of the envelope glycoproteins E1 and E2 with cell surface glycosaminoglycans. In this study, a variety of glycosaminoglycans, heparan sulfate (HS) from various bovine tissues as well as chondroitin sulfate (CS)/dermatan sulfate from bovine liver, were used to examine the direct interaction with recombinant E1 and E2 proteins. Intriguingly, among HS preparations from various bovine tissues, only liver HS strongly bound to both E1 and E2. Since HS from liver, which is the target tissue of HCV, contains highly sulfated structures compared to HS from other tissues, the present results suggest that HS-proteoglycan on the liver cell surface appears to be one of the molecules that define the liver-specific tissue tropism of HCV infection. The interaction assay with chemically modified heparin derivatives provided evidence that the binding of the viral proteins to heparin/HS is not only mediated by simple ionic interactions, but that the 6-O-sulfation and N-sulfation are important. Heparin oligosaccharides equal to or larger than 10-mer were required to inhibit the binding. Notably, a highly sulfated CS-E preparation from squid cartilage also strongly interacted with both viral proteins and inhibited the entry of pseudotype HCV into the target cells, suggesting that the highly sulfated CS-E might be useful as an anti-HCV drug.

Functional changes in the temporomandibular joint mechanoreceptors associated with experimentally induced condylar resorption in rats.

OBJECTIVES: To evaluate the influence of experimentally induced progressive condylar resorption (PCR) on temporomandibular joint (TMJ) mechanoreception. MATERIALS AND METHODS: Twenty 13-week-old male albino Wistar rats were divided equally into control and PCR groups. A compressive force was loaded on the left TMJ of PCR group rats to induce condylar resorption. Single-unit activities of TMJ mechanoreceptors were also induced through passive jaw movement. Recording was performed for the left Gasserian ganglion at 3 days and 1 week after the establishment of PCR group. The effects of PCR on TMJ units were assessed by measuring the firing threshold, maximum instantaneous firing frequency, and average firing frequency. RESULTS: Compared with the control group, there were no significant differences in the firing threshold of the PCR group after 3 days. The thresholds were significantly higher 1 week after compressive force loading on the condyle. The maximum instantaneous firing frequencies and the average firing frequencies showed no significant differences after 3 days. However, these were significantly lower 1 week after compressive force loading. CONCLUSIONS: The findings suggest that compressive force loading on the condyle may influence the function of TMJ mechanoreceptors.

Optimized Periodontal Regeneration for Orthodontics (O-PRO): A Case Series.

The fusion of orthodontic treatment and periodontal tissue-regeneration therapy has attracted attention. However, regenerated bone has a higher density than physiologic bone, which may cause problems including root resorption or stagnation of orthodontic movement. Therefore, the optimized periodontal regeneration for orthodontic movement (O-PRO) approach was developed with the aim of regenerating periodontal tissues with sparse bone quality. Unlike conventional methods, this concept is specifically suited for orthodontic movement. A new classification for the preoperative evaluation of periodontal tissues was also devised, and results are reported from cases where orthodontic treatment was implemented using each type of O-PRO method.

Nik-related kinase is targeted for proteasomal degradation by the chaperone-dependent ubiquitin ligase CHIP.

Nik-related kinase (Nrk) is a member of the germinal center kinase IV family and suppresses Akt signaling. In vivo, Nrk prevents placental hyperplasia and breast cancer formation. Here, we show that Nrk is regulated by the chaperone-dependent ubiquitin ligase carboxyl terminus of heat-shock protein (Hsp)70-interacting protein (CHIP). Immunoprecipitation and liquid chromatography-tandem mass spectrometry analysis reveal that Nrk preferentially interacts with CHIP and Hsp70/90 family proteins. Nrk protein levels are decreased by CHIP overexpression and increased by siRNA-mediated CHIP knockdown. Our results indicate that Nrk is ubiquitinated by CHIP in a chaperone-dependent manner, resulting in its proteasomal degradation. CHIP targets a fraction of Nrk molecules that have lost the ability to regulate Akt signaling. We conclude that CHIP plays an important role in regulating Nrk protein levels.

Functional adaptability of temporomandibular joint mechanoreceptors after an increase in the occlusal vertical dimension in rats.

OBJECTIVE: To investigate the effects of an experimentally-induced increase in the occlusal vertical dimension (iOVD) on the functional characteristics of temporomandibular joint (TMJ) mechanoreceptors in rats. MATERIALS AND METHODS: Sixty 13-week-old male albino Wistar rats were divided into control and iOVD groups (30 animals each). The vertical dimension between the maxillary and mandibular molars in the iOVD group was increased by 2.0 mm with a build-up of resin on the maxillary molars. Single-unit activities of TMJ mechanoreceptors were evoked by passive jaw movement. Recording was performed from the gasserian ganglion 1 day and 1, 3, 5, 7, and 9 weeks after the establishment of iOVD. RESULTS: Compared with the control group, the firing threshold was significantly lower at 1, 3, and 5 weeks after iOVD in the iOVD group. There were no significant differences in the firing threshold at 1 day, or 7 or 9 weeks. The maximum instantaneous firing frequency was significantly higher at 1, 3, and 5 weeks after iOVD in the iOVD group, but there were no significant differences at 1 day, or 7 or 9 weeks. There were no significant differences in the average firing frequency during the experimental period. CONCLUSIONS: The present study findings suggest that TMJ mechanoreceptors in adult rats may ultimately adapt to iOVD.

Chondroitin sulfate characterized by the E-disaccharide unit is a potent inhibitor of herpes simplex virus infectivity and provides the virus binding sites on gro2C cells.

Although cell surface chondroitin sulfate (CS) is regarded as an auxiliary receptor for binding of herpes simplex virus to cells, and purified CS chain types A, B, and C are known to interfere poorly or not at all with the virus infection of cells, we have found that CS type E (CS-E), derived from squid cartilage, exhibited potent antiviral activity. The IC(50) values ranged from 0.06 to 0.2 mug/ml and substantially exceeded the antiviral potency of heparin, the known inhibitor of virus binding to cells. Furthermore, in mutant gro2C cells that express CS but not heparan sulfate, CS-E showed unusually high anti-herpes virus activity with IC(50) values of <1 ng/ml. Enzymatic degradation of CS-E with chondroitinase ABC abolished its antiviral activity. CS-E inhibited the binding to cells of the purified virus attachment protein gC. A direct interaction of gC with immobilized CS-E and inhibition of this binding by CS-E oligosaccharide fragments greater than octasaccharide were demonstrated. Likewise, the gro2C-specific CS chains interfered with the binding of viral gC to these cells and were found to contain a considerable proportion (13%) of the E-disaccharide unit, suggesting that this unit is an essential component of the CS receptor for herpes simplex virus on gro2C cells and that the antiviral activity of CS-E was due to interference with the binding of viral gC to a CS-E-like receptor on the cell surface. Knowledge of the determinants of antiviral properties of CS-E will help in the development of inhibitors of herpes simplex virus infections in humans.