Engineering More Stable,Selectable Marker-Free Autoluminescent Mycobacteria by One Step

In our previous study, we demonstrated that the use of the autoluminescent Mycobacterium tuberculosis as a reporter strain had the potential to drastically reduce the time, effort, animals and costs consumed in evaluation of the activities of drugs and vaccines in live mice. However, the strains were relatively unstable and lost reporter with time without selection. The kanamycin selection marker used wasn’t the best choice as it provides resistance to amino glycosides which are an important class of second line drugs used in tuberculosis treatment. In addition, the marker could limit utility of the strains for screening of new potential drugs or evaluating drug combinations for tuberculosis treatment. Limited selection marker genes for mycobacterial genetic manipulation is a major drawback for such a marker-containing strain in many research fields. Therefore, selectable marker-free, more stable autoluminescent mycobacteria are highly needed. After trying several strategies, we created such mycobacterial strains successfully by using an integrative vector and removing both the resistance maker and integrase genes by Xer site-specific recombination in one step. The corresponding plasmid vectors developed in this study could be very convenient in constructing other selectable marker-free, more stable reporter mycobacteria with diverse applications.     

Overexpression of long non-coding RNA RP11-363E7.4 inhibits proliferation and invasion in gastric cancer

LncRNA RP11-363E7.4 has been shown to be downregulated in gastric cancer (GC), while the effect of lncRNA RP11-363E7.4 on GC and its potential molecular mechanisms is unclear. The purpose of this study was to explore the functional role and underlying molecular mechanisms of lncRNA RP11-363E7.4 involved in GC progress.To address the question, quantitative real-time PCR assay was performed to confirm lncRNA RP11-363E7.4 expression levels in GC tissues and cell lines. Cell proliferation, apoptosis, migration and invasion were estimated using Cell Counting Kit-8, colony formation, scratch wound healing and Transwell assays. Potential molecular mechanisms were evaluated using western blot assay. The results showed that lncRNA RP11-363E7.4 was significantly downregulated in GC cell lines and 82 paired tissues. The correlation between expression and clinicopathological features indicated that low expression of lncRNA RP11-363E7.4 was associated with T stage (P = .010). Functional experiments showed that overexpression of lncRNA RP11-363E7.4 prevented proliferation, migration, and invasion and induced apoptosis of GC cells. Western blot assay revealed that lncRNA RP11-363E7.4 functioned via the p53, Bax/Bcl-2, β-catenin pathway. In summary, this study revealed that lncRNA RP11-363E7.4 functioned as a tumour suppressor by inhibiting proliferation, migration, and invasion and inducing apoptosis of GC cells. Significance of the study :LncRNA RP11-363E7.4 has been shown to be downregulated in GC, while the effect of lncRNA RP11-363E7.4 on GC and its potential molecular mechanism is unclear. We revealed that lncRNA RP11-363E7.4 functioned as a tumour suppressor by inhibiting proliferation, migration, and invasion and inducing apoptosis of GC cells. LncRNA RP11-363E7.4 might become an attractive diagnostic and prognostic biomarker of GC and a promising target for GC treatment.     

PKD3 deficiency causes alterations in microtubule dynamics during the cell cycle

Protein kinase D 3 (PKD3) is a member of the PKD family that has been linked to many intracellular signaling pathways. However, defined statements regarding isoform specificity and in vivo functions are rare. Here, we use mouse embryonic fibroblast cells that are genetically depleted of PKD3 to identify isoform-specific functions. We show that PKD3 is involved in the regulation of the cell cycle by modulating microtubule nucleation and dynamics. In addition we also show that PKD1 partially can compensate for PKD3 function. Taken together our data provide new insights of a specific PKD3 signaling pathway by identifying a new function, which has not been identified before.     

Two-Way Horizontal and Vertical Omics Integration for Disease Subtype Discovery

Disease subtype discovery is an essential step in delivering personalized medicine. Disease subtyping via omics data has become a common approach for this purpose. With the advancement of technology and the lower price for generating omics data, multi-level and multi-cohort omics data are prevalent in the public domain, providing unprecedented opportunities to decrypt disease mechanisms. How to fully utilize multi-level/multi-cohort omics data and incorporate established biological knowledge toward disease subtyping remains a challenging problem. In this paper, we propose a meta-analytic integrative sparse Kmeans (MISKmeans) algorithm for integrating multi-cohort/multi-level omics data and prior biological knowledge. Compared with previous methods, MISKmeans shows better clustering accuracy and feature selection relevancy. An efficient R package, “MIS-Kmeans”, calling C++ is freely available on GitHub (https://github.com/Caleb-Huo/MIS-Kmeans).