Spatio-temporal association between mTOR and autophagy during cellular senescence

Evidence for a connection between lysosomes and mTOR is emerging. Seminal work from the Sabatini laboratory has shown that mTOR can be recruited to the lysosomal surface in response to amino acids, in a Rag GTPase-dependent manner, to become activated by Rheb. However the biological significance of this is not fully understood. Recent work from our laboratory has shown that lysosomes spatially link mTOR and autophagy forming a cytoplasmic compartment in close proximity to the Golgi apparatus (GA) during oncogenic Ras-induced senescence. The TOR-autophagy spatial coupling compartment (TASCC) is enriched for autolysosomes, but largely excludes autophagosomes. Our data suggest that mTOR, which is a positive regulator of protein synthesis, is recruited, in part, by the amino acid-rich environment surrounding the autolysosomes. This then facilitates protein synthesis at the nearby rER-GA system, reinforcing lysosome and autophagy biogenesis. Proper TASCC formation contributes to the production of secretory proteins, which also utilizes the rER-GA system. Since mTOR inhibits autophagy during the initial stages of autophagosome formation, TASCC formation is likely to facilitate autophagy by sequestering mTOR, suggesting that the TASCC is a self-enhancing structure.     

Extensive Chromosome Homoeology among Brassiceae Species Were Revealed by Comparative Genetic Mapping with High-Density EST-Based SNP Markers in Radish (Raphanus sativus L.)

A linkage map of expressed sequence tag (EST)-based markers in radish (Raphanus sativus L.) was constructed using a low-cost and high-efficiency single-nucleotide polymorphism (SNP) genotyping method named multiplex polymerase chain reaction–mixed probe dot-blot analysis developed in this study. Seven hundred and forty-six SNP markers derived from EST sequences of R. sativus were assigned to nine linkage groups with a total length of 806.7 cM. By BLASTN, 726 markers were found to have homologous genes in Arabidopsis thaliana, and 72 syntenic regions, which have great potential for utilizing genomic information of the model species A. thaliana in basic and applied genetics of R. sativus, were identified. By construction and analysis of the genome structures of R. sativus based on the 24 genomic blocks within the Brassicaceae ancestral karyotype, 23 of the 24 genomic blocks were detected in the genome of R. sativus, and half of them were found to be triplicated. Comparison of the genome structure of R. sativus with those of the A, B, and C genomes of Brassica species and that of Sinapis alba L. revealed extensive chromosome homoeology among Brassiceae species, which would facilitate transfer of the genomic information from one Brassiceae species to another.     

AHR,a novel acute hypoxia‐response sequence,drives reporter gene expression under hypoxia in vitro and in vivo

ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin motifs 1) is an early immediate gene. We have previously reported that ADAMTS1 was strongly induced by hypoxia. In this study, we investigated whether ADAMTS1 promoter‐driven reporter signal is detectable by acute hypoxia. We constructed the GFP (green fluorescent protein) expression vector [AHR (acute hypoxia‐response sequence)‐GFP] under the control of ADAMTS1 promoter and compared it with the constitutive GFP‐expressing vector under the control of CMV (cytomegalovirus promoter‐GFP). We transduced AHR‐GFP and examined whether GFP signals can be detected under the acute hypoxia. When the human umbilical vein [HUVEC (human umbilical vein endothelial cells)] was transduced under normoxia, there were few GFP signals, while CMV‐GFP showed considerable GFP signals. When HUVEC was stimulated with hypoxia, GFP signals from AHR‐GFP gene were induced under hypoxic conditions. Notably, the GFP signals peaked at 3 h under hypoxia. In ischaemic hind limb model, transduced AHR‐GFP showed hypoxic induction of GFP signals. In summary, we have demonstrated that the AHR system induced the reporter gene expression by acute hypoxia, and its induction is transient. This is the first report showing the unique acute hypoxia‐activated gene expression system.     

Geographical distribution pattern and interisland movements of Orii’s flying fox in Okinawa Islands, the Ryukyu Archipelago, Japan

The study of mobile animals such as flying foxes in insular habitats involves clarifying the population status on each island and determining the factors affecting movement patterns among the islands in their distributional range. We visited 25 of the Okinawa Islands and documented the number of Orii’s flying foxes Pteropus dasymallus inopinatus from August 2005 to May 2006. We also conducted a monthly road census on the main island (Okinawa-jima Island) and six adjacent islands from June 2006 to January 2007 and counted the number of fruit-bearing trees of the bats’ four main food plants. The results of classification and regression tree analysis suggested that distance from the main island was a primary factor in determining the distribution pattern and population size of this flying fox, whereas island area, number of plant species, and food availability did not directly affect population size. The number of flying foxes on each island tended to decrease with an increase in distance from the main island; no flying foxes existed on islands >30 km away from the main island. On the other hand, the results of the monthly census showed that the population size on each island fluctuated seasonally. Individuals may move between islands in response to seasonal changes in food availability. In conclusion, the distribution and abundance of Orii’s flying foxes in the Okinawa Islands may be determined by the rate of immigration/emigration, depending on each island’s distance from the main island. Seasonal changes in food availability may act as a trigger for interisland movement, but that movement may be restricted by island connectivity.     

High sero-prevalence of caseous lymphadenitis identified in slaughterhouse samples as a consequence of deficiencies in sheep farm management in the state of Minas Gerais, Brazil

Background

Multiple congenital ocular anomalies (MCOA) syndrome is a hereditary congenital eye defect that was first described in Silver colored Rocky Mountain horses. The mutation causing this disease is located within a defined chromosomal interval, which also contains the gene and mutation that is associated with the Silver coat color (PMEL17, exon 11). Horses that are homozygous for the disease-causing allele have multiple defects (MCOA-phenotype), whilst the heterozygous horses predominantly have cysts of the iris, ciliary body or retina (Cyst-phenotype). It has been argued that these ocular defects are caused by a recent mutation that is restricted to horses that are related to the Rocky Mountain Horse breed. For that reason we have examined another horse breed, the Icelandic horse, which is historically quite divergent from Rocky Mountain horses.

Results

We examined 24 Icelandic horses and established that the MCOA syndrome is present in this breed. Four of these horses were categorised as having the MCOA-phenotype and were genotyped as being homozygous for the PMEL17 mutation. The most common clinical signs included megaloglobus, iris stromal hypoplasia, abnormal pectinate ligaments, iridociliary cysts occasionally extending into the peripheral retina and cataracts. The cysts and pectinate ligament abnormalities were observed in the temporal quadrant of the eyes. Fourteen horses were heterozygous for the PMEL17 mutation and were characterized as having the Cyst-phenotype with cysts and occasionally curvilinear streaks in the peripheral retina. Three additional horses were genotyped as PMEL17 heterozygotes, but in these horses we were unable to detect cysts or other forms of anomalies. One eye of a severely vision-impaired 18 month-old stallion, homozygous for the PMEL17 mutation was examined by light microscopy. Redundant duplication of non-pigmented ciliary body epithelium, sometimes forming cysts bulging into the posterior chamber and localized areas of atrophy in the peripheral retina were seen.

Conclusions

The MCOA syndrome is segregating with the PMEL17 mutation in the Icelandic Horse population. This needs to be taken into consideration in breeding decisions and highlights the fact that MCOA syndrome is present in a breed that are more ancient and not closely related to the Rocky Mountain Horse breed.     

Combination of hTERT and bmi-1, E6, or E7 induces prolongation of the life span of bone marrow stromal cells from an elderly donor without affecting their neurogenic potential

Murine bone marrow stromal cells differentiate not only into mesodermal derivatives, such as osteocytes, chondrocytes, adipocytes, skeletal myocytes, and cardiomyocytes, but also into neuroectodermal cells in vitro. Human bone marrow stromal cells are easy to isolate but difficult to study because of their limited life span. To overcome this problem, we attempted to prolong the life span of bone marrow stromal cells and investigated whether bone marrow stromal cells modified with bmi-1, hTERT, E6, and E7 retained their differentiated capability, or multipotency. In this study, we demonstrated that the life span of bone marrow stromal cells derived from a 91-year-old donor could be extended and that the stromal cells with an extended life span differentiated into neuronal cells in vitro. We examined the neuronally differentiated cells morphologically, physiologically, and biologically and compared the gene profiles of undifferentiated and differentiated cells. The neuronally differentiated cells exhibited characteristics similar to those of midbrain neuronal progenitors. Thus, the results of this study support the possible use of autologous-cell graft systems to treat central nervous system diseases in geriatric patients.