Silencing of FABP3 Inhibits Proliferation and Promotes Apoptosis in Embryonic Carcinoma Cells

Fatty acid–binding protein 3 (FABP3) facilitates the movement of fatty acids in cardiac muscle. Previously, we reported that FABP3 is highly upregulated in the myocardium of ventricular septal defect patients and overexpression of FABP3 inhibited proliferation and promoted apoptosis in embryonic carcinoma cells (P19 cells). In this study, we aimed to investigate the effect of FABP3 gene silencing on P19 cell differentiation, proliferation and apoptosis. We used RNA interference and a lentiviral-based vector system to create a stable FABP3-silenced P19 cell line; knockdown of FABP3 was confirmed by quantitative real-time PCR. Expression analysis of specific differentiation marker genes using quantitative real-time PCR and observation of morphological changes using an inverted microscope revealed that knockdown of FABP3 did not significantly affect the differentiation of P19 cells into cardiomyocytes. CCK-8 proliferation assays and cell cycle analysis demonstrated that FABP3 gene silencing significantly inhibited P19 cell proliferation. Furthermore, Annexin V-FITC/propidium iodide staining and the caspase-3 activity assay revealed that FABP3 gene silencing significantly promoted serum starvation–induced apoptosis in P19 cells. In agreement with our previous research, these results demonstrate that FABP3 may play an important role during embryonic heart development, and that either overexpression or silencing of FABP3 will lead to an imbalance between proliferation and apoptosis, which may result in embryonic cardiac malformations.     

T Cell-Specific Overexpression of TGF?1 Fails to Influence Atherosclerosis in ApoE-Deficient Mice

Clinical data have indicated a negative correlation between plasma TGFß1 concentrations and the extent of atherosclerosis and have thus led to the hypothesis that the pleiotropic cytokine may have anti-atherogenic properties. T-cells are currently discussed to significantly participate in atherogenesis, but the precise role of adaptive immunity in atherogenesis remains to be elucidated. TGFß1 is known to strongly modulate the function of T-cells, however, inhibition of TGFß1 signalling in T-cells of atherosclerosis-prone knock-out mice failed to unequivocally clarify the role of the cytokine for the development of atherosclerosis. In the present study, we thus tried to specify the role of TGFß1 in atherogenesis by using the murine CD2-TGFß1 transgenic strain which represents a well characterized model of T-cell specific TGFß1 overexpression. The CD2-TGFß1 transgenic mice were crossed to ApoE knock-out mice and quantity and quality of atherosclerosis regarding number of macrophages, smooth muscle cells, CD3 positive T-cells and collagen was analyzed in CD2-TGFß1 ApoE double mutants as well as non-transgenic ApoE controls on both normal and atherogenic diet of a duration of 8, 16 or 24 weeks, respectively. In all experimental groups investigated, we failed to detect any influence of TGFß1 overexpression on disease. Total number of CD3-positive T-lymphocytes was not significantly different in atherosclerotic lesions of CD2-TGFß1 ApoE^−/− females and isogenic ApoE^−/− controls, even after 24 weeks on the atherogenic diet. The synopsis of these data and our previous study on TGFß1 overexpressing macrophages suggests that potential effects of TGFß1 on atherosclerosis are most probably mediated by macrophages rather than T-cells.     

Simulation of the Planetary Interior Differentiation Processes in the Laboratory

A planetary interior is under high-pressure and high-temperature conditions and it has a layered structure. There are two important processes that led to that layered structure, (1) percolation of liquid metal in a solid silicate matrix by planet differentiation, and (2) inner core crystallization by subsequent planet cooling. We conduct high-pressure and high-temperature experiments to simulate both processes in the laboratory. Formation of percolative planetary core depends on the efficiency of melt percolation, which is controlled by the dihedral (wetting) angle. The percolation simulation includes heating the sample at high pressure to a target temperature at which iron-sulfur alloy is molten while the silicate remains solid, and then determining the true dihedral angle to evaluate the style of liquid migration in a crystalline matrix by 3D visualization. The 3D volume rendering is achieved by slicing the recovered sample with a focused ion beam (FIB) and taking SEM image of each slice with a FIB/SEM crossbeam instrument. The second set of experiments is designed to understand the inner core crystallization and element distribution between the liquid outer core and solid inner core by determining the melting temperature and element partitioning at high pressure. The melting experiments are conducted in the multi-anvil apparatus up to 27 GPa and extended to higher pressure in the diamond-anvil cell with laser-heating. We have developed techniques to recover small heated samples by precision FIB milling and obtain high-resolution images of the laser-heated spot that show melting texture at high pressure. By analyzing the chemical compositions of the coexisting liquid and solid phases, we precisely determine the liquidus curve, providing necessary data to understand the inner core crystallization process.     

The History and Advances of Reversible Terminators Used in New Generations of Sequencing Technology

DNA sequencing using reversible terminators, as one sequencing by synthesis strategy, has garnered a great deal of interest due to its popular application in the second-generation high-throughput DNA sequencing technology. In this review, we provided its history of development, classification, and working mechanism of this technology. We also outlined the screening strategies for DNA polymerases to accommodate the reversible terminators as substrates during polymerization; particularly, we introduced the "REAP" method developed by us. At the end of this review, we discussed current limitations of this approach and provided potential solutions to extend its application.     

Proteomic methods reveal cyclophilin a function as a host restriction factor against rotavirus infection

Rotavirus (RV) infection is the main cause of acute dehydrating diarrhea in infants and young children below 5 years old worldwide. RV infection causes a global shutoff of host proteins as many other viruses do. However, previous studies revealed that RV could selectively upregulated the expression of some host proteins that then played important roles in RV infection. To globally explor such host proteins that were upregulated in early human rotavirus (HRV) infection, proteomic methods were used and a total of ten upregulated host proteins were unambiguously identified. Cyclophilin A (CYPA), a peptidyl‐prolyl cis‐trans isomerase, was among these upregulated host proteins. Following infection, CYPA was recruited to the viroplasm and interacted with HRV structural protein VP2; CYPA reduced host susceptibility to HRV infection and inhibited replication of HRV by repressing the expression of viral proteins. Furthermore, we found that the increased expression of CYPA in enterocytes of small intestine correlated to the period when BALB/c mice became resistant to RV diarrhea. Together, we identified CYPA as a novel host restriction factor that confered protection against RV infection and might contribute to host susceptibility to RV diarrhea.     

SLC26A4 Targeted to the Endolymphatic Sac Rescues Hearing and Balance in Slc26a4 Mutant Mice

Mutations of SLC26A4 are a common cause of human hearing loss associated with enlargement of the vestibular aqueduct. SLC26A4 encodes pendrin, an anion exchanger expressed in a variety of epithelial cells in the cochlea, the vestibular labyrinth and the endolymphatic sac. Slc26a4 ^Δ/Δ mice are devoid of pendrin and develop a severe enlargement of the membranous labyrinth, fail to acquire hearing and balance, and thereby provide a model for the human phenotype. Here, we generated a transgenic mouse line that expresses human SLC26A4 controlled by the promoter of ATP6V1B1. Crossing this transgene into the Slc26a4 ^Δ/Δ line restored protein expression of pendrin in the endolymphatic sac without inducing detectable expression in the cochlea or the vestibular sensory organs. The transgene prevented abnormal enlargement of the membranous labyrinth, restored a normal endocochlear potential, normal pH gradients between endolymph and perilymph in the cochlea, normal otoconia formation in the vestibular labyrinth and normal sensory functions of hearing and balance. Our study demonstrates that restoration of pendrin to the endolymphatic sac is sufficient to restore normal inner ear function. This finding in conjunction with our previous report that pendrin expression is required for embryonic development but not for the maintenance of hearing opens the prospect that a spatially and temporally limited therapy will restore normal hearing in human patients carrying a variety of mutations of SLC26A4.     

GsSnRK1 interplays with transcription factor GsERF7 from wild soybean to regulate soybean stress resistance

Although the function and regulation of SnRK1 have been studied in various plants, its molecular mechanisms in response to abiotic stresses are still elusive. In this work, we identified an AP2/ERF domain-containing protein (designated GsERF7) interacting with GsSnRK1 from a wild soybean cDNA library. GsERF7 gene expressed dominantly in wild soybean roots and was responsive to ethylene, salt, and alkaline. GsERF7 bound GCC cis-acting element and could be phosphorylated on S36 by GsSnRK1. GsERF7 phosphorylation facilitated its translocation from cytoplasm to nucleus and enhanced its transactivation activity. When coexpressed in the hairy roots of soybean seedlings, GsSnRK1(wt) and GsERF7(wt) promoted plants to generate higher tolerance to salt and alkaline stresses than their mutated species, suggesting that GsSnRK1 may function as a biochemical and genetic upstream kinase of GsERF7 to regulate plant adaptation to environmental stresses. Furthermore, the altered expression patterns of representative abiotic stress-responsive and hormone-synthetic genes were determined in transgenic soybean hairy roots after stress treatments. These results will aid our understanding of molecular mechanism of how SnRK1 kinase plays a cardinal role in regulating plant stress resistances through activating the biological functions of downstream factors.     

Screening of optimal reference genes for qRT-PCR and preliminary exploration of cold resistance mechanisms in Prunus mume and Prunus sibirica varieties

Molecular Biology Reports - Prunus sibirica and Prunus mume are closely related plant species that differ in cold tolerance. Hybrids of P. sibirica and true mume, belonging to the apricot mei...