A non-canonical function of Plk4 in centriolar satellite integrity and ciliogenesis through PCM1 phosphorylation

Centrioles are the major constituents of the animal centrosome, in which Plk4 kinase serves as a master regulator of the duplication cycle. Many eukaryotes also contain numerous peripheral particles known as centriolar satellites. While centriolar satellites aid centriole assembly and primary cilium formation, it is unknown whether Plk4 plays any regulatory roles in centriolar satellite integrity. Here we show that Plk4 is a critical determinant of centriolar satellite organisation. Plk4 depletion leads to the dispersion of centriolar satellites and perturbed ciliogenesis. Plk4 interacts with the satellite component PCM1, and its kinase activity is required for phosphorylation of the conserved S372. The nonphosphorylatable PCM1 mutant recapitulates phenotypes of Plk4 depletion, while the phosphomimetic mutant partially rescues the dispersed centriolar satellite patterns and ciliogenesis in cells depleted of PCM1. We show that S372 phosphorylation occurs during the G1 phase of the cell cycle and is important for PCM1 dimerisation and interaction with other satellite components. Our findings reveal that Plk4 is required for centriolar satellite function, which may underlie the ciliogenesis defects caused by Plk4 dysfunction

Altered Homeostasis of CD4+ Memory T Cells in Allogeneic Hematopoietic Stem Cell Transplant Recipients: Chronic Graft-versus-Host Disease Enhances T Cell Differentiation and Exhausts Central Memory T Cell Pool

AbstractAn increased risk of late infection is a serious complication after allogeneic hematopoietic stem cell transplantation (AHSCT), especially for recipients with defective CD4+ T cell recovery. Although chronic graft-versus-host disease (cGVHD) negatively influences CD4+ T cell reconstitution, the mechanisms leading to this defect are not well understood. We found that the proportion of CD27− CD4+ T cells was remarkably increased in ASHCT recipients with cGVHD or with repetitive infectious episodes. Isolated CD27− CD4+ T cells from ASHCT recipients had significantly shortened telomere length, displayed enhanced vulnerability to activation-induced cell death, and showed extremely reduced clonal diversity, when compared with CD27− CD4+ T cells from healthy donors. Also, CD27+ CD4+ T cells from AHSCT recipients easily lost their expression of CD27 in response to antigen stimulation regardless of cGVHD status. Taken together, these data indicate that homeostasis of memory CD4+ T cells from AHSCT recipients is altered, and that they easily transit into CD27− effector memory T cells. Increased in vivo T cell stimulation observed in recipients with cGVHD further promotes the transition to effector memory cells, a change that decreases the central memory CD4+ T cell pool and consequently weakens the recipient’s defense against persistently infecting pathogens

The conserved Wdr8-hMsd1/SSX2IP complex localises to the centrosome and ensures proper spindle length and orientation

The centrosome plays a pivotal role in a wide range of cellular processes and its dysfunction is causally linked to many human diseases including cancer and developmental and neurological disorders. This organelle contains more than one hundred components, and yet many of them remain uncharacterised. Here we identified a novel centrosome protein Wdr8, based upon the structural conservation of the fission yeast counterpart. We showed that Wdr8 constitutively localises to the centrosome and super resolution microscopy uncovered that this protein is enriched at the proximal end of the mother centriole. Furthermore, we identified hMsd1/SSX2IP, a conserved spindle anchoring protein, as one of Wdr8 interactors by mass spectrometry. Wdr8 formed a complex and partially colocalised with hMsd1/SSX2IP. Intriguingly, knockdown of Wdr8 or hMsd1/SSX2IP displayed very similar mitotic defects, in which spindle microtubules became shortened and misoriented. Indeed, Wdr8 depletion resulted in the reduced recruitment of hMsd1/SSX2IP to the mitotic centrosome, though the converse is not true. Together, we propose that the conserved Wdr8-hMsd1/SSX2IP complex plays a critical role in controlling proper spindle length and orientation.T.T. and A.P.S were supported by Cancer Research UK.Supplementary data related to this article can be found at http://dx.doi.org/10.1016/j.bbrc.2015.10.169

Crosstalk between the Rb Pathway and AKT Signaling Forms a Quiescence-Senescence Switch

SummaryCell-cycle arrest in quiescence and senescence is largely orchestrated by the retinoblastoma (Rb) tumor-suppressor pathway, but the mechanisms underlying the quiescence-senescence switch remain unclear. Here, we show that the crosstalk between the Rb-AKT-signaling pathways forms this switch by controlling the overlapping functions of FoxO3a and FoxM1 transcription factors in cultured fibroblasts. In the absence of mitogenic signals, although FoxM1 expression is repressed by the Rb pathway, FoxO3a prevents reactive oxygen species (ROS) production by maintaining SOD2 expression, leading to quiescence. However, if the Rb pathway is activated in the presence of mitogenic signals, FoxO3a is also inactivated by AKT, thus reducing SOD2 expression and consequently allowing ROS production. This situation elicits senescence through irreparable DNA damage. We demonstrate that this pathway operates in mouse liver, indicating that this machinery may contribute more broadly to tissue homeostasis in vivo

Establishment and evaluation of the suspension culture system for umbilical cord- derived mesenchymal stromal cells

Mesenchymal stromal cells (MSCs) derived from various tissues including bone marrow, adipose and umbilical cord tissues have been shown to modulate aberrantly activated immune system. With the features, MSC-based therapies targeting graft-versus-host disease (GvHD) by the administration of bone marrow-derived MSCs (BM-MSCs) have been available in some countries including Japan, and the expectations for the stable and cost effective supply system are getting higher and higher recently. However, the conventional culture systems which usually use plastic flask or multi-chamber equipment require space and manpower, thus the maximal expansion of MSCs at one production is likely to be limited. To compensate the limitation, repetitive productions have been unavoidable, and higher the production cost. Here, we introduced a new suspension-culture system, using micro-carriers and single-use-bioreactors, for the preparation of MSCs in anticipation of establishment of mass production system. Since the umbilical cord (UC) tissues can be collected through noninvasive procedure, and UC-derived MSCs (UC-MSCs) are shown to present higher proliferation rate and lower immunogenicity in comparison with BM-MSCs, we evaluated the potential and the versatility of UC-MSCs for the treatment of several diseases including GvHD. Results from several in vitro assays demonstrated that our new culture system maintains major key characteristics of MSCs, such as adhesiveness to cell culture surface, the expression of cell surface markers, differentiation capacities toward osteoblasts, chondroblasts, and adipocytes, and immunosuppressive effects on activated T cells. We are currently investigating cellular profiles and characteristics which are specific to the cells prepared in our suspension-culture system through meta-analysis. The established suspension-culture system is presumed to attain the mass production of UC-MSCs, contributing to lower the cost and also providing possible applications for MSCs from other origins

In Situ Hybridization Analysis of the Expression of Futsch, Tau, and MESK2 Homologues in the Brain of the European Honeybee (Apis mellifera L.)

BACKGROUND: The importance of visual sense in Hymenopteran social behavior is suggested by the existence of a Hymenopteran insect-specific neural circuit related to visual processing and the fact that worker honeybee brain changes morphologically according to its foraging experience. To analyze molecular and neural bases that underlie the visual abilities of the honeybees, we used a cDNA microarray to search for gene(s) expressed in a neural cell-type preferential manner in a visual center of the honeybee brain, the optic lobes (OLs). METHODOLOGY/PRINCIPAL FINDINGS: Expression analysis of candidate genes using in situ hybridization revealed two genes expressed in a neural cell-type preferential manner in the OLs. One is a homologue of Drosophila futsch, which encodes a microtubule-associated protein and is preferentially expressed in the monopolar cells in the lamina of the OLs. The gene for another microtubule-associated protein, tau, which functionally overlaps with futsch, was also preferentially expressed in the monopolar cells, strongly suggesting the functional importance of these two microtubule-associated proteins in monopolar cells. The other gene encoded a homologue of Misexpression Suppressor of Dominant-negative Kinase Suppressor of Ras 2 (MESK2), which might activate Ras/MAPK-signaling in Drosophila. MESK2 was expressed preferentially in a subclass of neurons located in the ventral region between the lamina and medulla neuropil in the OLs, suggesting that this subclass is a novel OL neuron type characterized by MESK2-expression. These three genes exhibited similar expression patterns in the worker, drone, and queen brains, suggesting that they function similarly irrespective of the honeybee sex or caste. CONCLUSIONS: Here we identified genes that are expressed in a monopolar cell (Amfutsch and Amtau) or ventral medulla-preferential manner (AmMESK2) in insect OLs. These genes may aid in visualizing neurites of monopolar cells and ventral medulla cells, as well as in analyzing the function of these neurons

Serum 25-Hydroxyvitamin D Concentrations and Season-Specific Correlates in Japanese Adults

Background: Several lines of evidence indicate an important role for vitamin D in the prevention of a range of diseases. Blood vitamin D levels show clear seasonal variation; however, data on the determinants of vitamin D status for each season are limited. We investigated the association between lifestyle and serum vitamin D concentration by season in Japanese workers. Methods: Subjects were 312 men and 217 women aged 21 to 67 years who worked in municipal offices in Northern Kyushu, Japan and participated in a periodic checkup in July or November. Multiple linear regression analysis was used to examine the association between serum 25-hydroxivitamin D concentrations and lifestyle factors for each season. Results: Mean serum 25-hydroxyvitamin D concentration was 27.4 ng/ml (68.4 nmol/L) and 21.4 ng/ml (53.4 nmol/L) for workers surveyed in July and November, respectively (P < 0.001); the prevalence of vitamin D deficiency (<20 ng/ml) was 9.3% and 46.7%, respectively (P < 0.001). In November, dietary vitamin D intake (in both sexes) and nonsmoking and physical activity (in men) were significantly associated with higher concentrations of serum 25-hydroxyvitamin D. In summer, fish/shellfish intake was associated with higher serum 25- hydroxyvitamin D concentrations in women. Conclusions: Vitamin D deficiency is common in Japanese workers during seasons with limited sunlight. The lifestyle correlates of favorable vitamin D status in November were physical activity, dietary vitamin D intake, and nonsmoking