Modulating signaling networks by CRISPR/Cas9-mediated transposable element insertion

In a recent past, transposable elements (TEs) were referred to as selfish genetic components only capable of copying themselves with the aim of increasing the odds of being inherited. Nonetheless, TEs have been initially proposed as positive control elements acting in synergy with the host. Nowadays, it is well known that TE movement into host genome comprises an important evolutionary mechanism capable of increasing the adaptive fitness. As insights into TE functioning are increasing day to day, the manipulation of transposition has raised an interesting possibility of setting the host functions, although the lack of appropriate genome engineering tools has unpaved it. Fortunately, the emergence of genome editing technologies based on programmable nucleases, and especially the arrival of a multipurpose RNA-guided Cas9 endonuclease system, has made it possible to reconsider this challenge. For such purpose, a particular type of transposons referred to as miniature inverted-repeat transposable elements (MITEs) has shown a series of interesting characteristics for designing functional drivers. Here, recent insights into MITE elements and versatile RNA-guided CRISPR/Cas9 genome engineering system are given to understand how to deploy the potential of TEs for control of the host transcriptional activity.Fil: Vaschetto, Luis Maria Benjamin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Diversidad y Ecología Animal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto de Diversidad y Ecología Animal; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Cátedra de Diversidad Animal I; Argentin

Analysis for genotyping Duffy blood group in inhabitants of Sudan, the Fourth Cataract of the Nile

<p>Abstract</p> <p>Background</p> <p>Genetic polymophisms of the Duffy antigen receptor for the chemokines (DARC) gene successfully protected against blood stage infection by <it>Plasmodium vivax </it>infection. The Fy (a-, b-) phenotype is predominant among African populations, particularly those originating from West Africa, and it is rare among non-African populations. The aim of this study was to analyse the frequency of four Duffy blood groups based on SNPs (T-33C, G125A, G298A and C5411T) in two local tribes of Sudanese Arabs, the <it>Shagia </it>and <it>Manasir</it>, which are both from the region of the Fourth Nile cataract in Sudan.</p> <p>Methods</p> <p>An analysis of polymorphisms was performed on 217 individuals (126 representatives of the <it>Shagia </it>tribe and 91 of the <it>Manasir)</it>. Real-time PCR and TaqMan Genotyping Assays were used to study the prevalence of alleles and genotypes.</p> <p>Results</p> <p>The analysis of allelic and genotype frequency in the T-33C polymorphisms demonstrated a significant dominance of the <it>C </it>allele and <it>CC </it>genotype (OR = 0.53 [0.32-0.88]; p = 0.02) in both tribes. The G125A polymorphism is associated with phenotype Fy(a-, b-) and was identified in 83% of <it>Shagia </it>and 77% of <it>Manasir</it>. With regard to G298A polymorphisms, the genotype frequencies were different between the tribes (p = 0,002) and no single <it>AA </it>homozygote was found. Based on four SNPs examined, 20 combinations of genotypes for the <it>Shagia </it>and <it>Manasir </it>tribes were determined. The genotype <it>CC/AA/GG/CT </it>occurred most often in <it>Shagia </it>tribe (45.9%) but was rare in the <it>Manasir </it>tribe (6.6%) (p < 0.001 <it>Shagia </it>versus <it>Manasir</it>). The <it>FY*A^ES</it>allele was identified in both analysed tribes. The presence of individuals with the <it>FY*A/FY*A </it>genotype was demonstrated only in the <it>Shagia </it>tribe.</p> <p>Conclusion</p> <p>This is probably the first report showing genotypically Duffy-negative people who carry both <it>FY*B^ES</it>and <it>FY*A^ES</it>. The identification of the <it>FY*A^ES</it>allele in both tribes may be due to admixture of the non-African genetic background. Taken as a whole, allele and genotype frequencies between the <it>Shagia </it>and the <it>Manasir </it>were statistically different. However, the presence of individuals with the <it>FY*A/FY*A </it>genotype was demonstrated only in the <it>Shagia </it>tribe.</p

Wnt/β-Catenin Signaling Induces the Aging of Mesenchymal Stem Cells through the DNA Damage Response and the p53/p21 Pathway

Recent studies have demonstrated the importance of cellular extrinsic factors in the aging of adult stem cells. However, the effects of an aged cell–extrinsic environment on mesenchymal stem cell (MSC) aging and the factors involved remain unclear. In the current study, we examine the effects of old rat serum (ORS) on the aging of MSCs, and explore the effects and mechanisms of Wnt/β-catenin signaling on MSC aging induced by ORS treatment. Senescence-associated changes in the cells are examined with SA-β-galactosidase staining and ROS staining. The proliferation ability is detected by MTT assay. The surviving and apoptotic cells are determined using AO/EB staining. The results suggest that ORS promotes MSC senescence and reduces the proliferation and survival of cells. The immunofluorescence staining shows that the expression of β-catenin increases in MSCs of old rats. To identify the effects of Wnt/β-catenin signaling on MSC aging induced with ORS, the expression of β-catenin, GSK-3β, and c-myc are detected. The results show that the Wnt/β-catenin signaling in the cells is activated after ORS treatment. Then we examine the aging, proliferation, and survival of MSCs after modulating Wnt/β-catenin signaling. The results indicate that the senescence and dysfunction of MSCs in the medium containing ORS is reversed by the Wnt/β-catenin signaling inhibitor DKK1 or by β-catenin siRNA. Moreover, the expression of γ-H2A.X, a molecular marker of DNA damage response, p16INK4a, p53, and p21 is increased in senescent MSCs induced with ORS, and is also reversed by DKK1 or by β-catenin siRNA. In summary, our study indicates the Wnt/β-catenin signaling may play a critical role in MSC aging induced by the serum of aged animals and suggests that the DNA damage response and p53/p21 pathway may be the main mediators of MSC aging induced by excessive activation of Wnt/β-catenin signaling

Leishmania-Specific Surface Antigens Show Sub-Genus Sequence Variation and Immune Recognition

Single-celled Leishmania parasites, transmitted by sand flies, infect humans and other mammals in many tropical and sub-tropical regions, giving rise to a spectrum of diseases called the leishmaniases. Species of parasite within the Leishmania genus can be divided into two groups (referred to as sub-genera) that are separated by up to 100 million years of evolution yet are highly related at the genome level. Our research is focused on identifying gene differences between these sub-genera that may identify proteins that impact on the transmission and pathogenicity of different Leishmania species. Here we report the presence of a highly-variant genomic locus (OHL) that was previously described as absent in parasites of the L. (Viannia) subgenus (on the basis of lack of key genes) but is present and well-characterised (as the LmcDNA16 locus) in all members of the alternative subgenus, L. (Leishmania). We demonstrate that the proteins encoded within the LmcDNA16 and OHL loci are similar in their structure and surface localisation in mammalian-infective amastigotes, despite significant differences in their DNA sequences. Most importantly, we demonstrate that the OHL locus proteins, like the HASP proteins from the LmcDNA16 locus, contain highly variable amino acid repeats that are antigenic in man and may therefore contribute to future vaccine development