Chemical composition, seasonal variation, and biosynthetic considerations of essential oils from Baccharis microdonta and B. elaeagnoides (Asteraceae)

The chemical composition and seasonal variation throughout one year of the essential oils from leaves of Baccharis microdonta and B. elaeagnoides, collected in Campos do Jordão, SP, were investigated. The composition of the latter species has been described for the first time. By GC (RI) and GC/MS analysis, 43 compounds were identified, and a predominance of oxygenated sesquiterpene derivatives was found in both species. The main components of the B. microdonta oils were elemol ( 31 ; 11.7–30.6%), spathulenol ( 34 ; 4.7–9.1%), β‐caryophyllene ( 19 ; 3.7–6.2%), and germacrene D ( 24 ; 2.9–12.2%), and those of the B. elaeagnoides oils were 34 (10.1–21.5%), viridiflorol ( 35 ; 3.6–18.4%), 24 (0.9–13.8%), and 19 (3.5–9.4%). The identified compounds were grouped according to their respective C‐skeletons, and the percentages of occurrence of the C‐skeletons in the essential oils of leaves collected in the four seasons allowed identifying the preferential accumulation of different types of C‐skeletons as well as the seasonal variation of the biosynthetic routes over the studied period.     

Proteome of Human Stem Cells from Periodontal Ligament and Dental Pulp

Background

Many adult tissues contain a population of stem cells with the ability to regenerate structures similar to the microenvironments from which they are derived in vivo and represent a promising therapy for the regeneration of complex tissues in the clinical disorder. Human adult stem cells (SCs) including bone marrow stem cells (BMSCs), dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs) have been characterized for their high proliferative potential, expression of characteristic SC-associated markers and for the plasticity to differentiate in different lineage in vitro.

Methodology/Principal Findings

The aim of this study is to define the molecular features of stem cells from oral tissue by comparing the proteomic profiles obtained with 2-DE followed by MALDI-TOF/TOF of ex-vivo cultured human PDLSCs, DPSCs and BMSCs. Our results showed qualitative similarities in the proteome profiles among the SCs examined including some significant quantitative differences. To enrich the knowledge of oral SCs proteome we performed an analysis in narrow range pH 4–7 and 6–9, and we found that DPSCs vs PDLSCs express differentially regulated proteins that are potentially related to growth, regulation and genesis of neuronal cells, suggesting that SCs derived from oral tissue source populations may possess the potential ability of neuronal differentiation which is very consistent with their neural crest origin.

Conclusion/Significance

This study identifies some differentially expressed proteins by using comparative analysis between DPSCs and PDLSCs and BMSCs and suggests that stem cells from oral tissue could have a different cell lineage potency compared to BMSCs.     

Monoclonal antibodies toward different Tn-amino acid backbones display distinct recognition patterns on human cancer cells. Implications for effective immuno-targeting of cancer

The Tn antigen (GalNAcα-O-Ser/Thr) is a well-established tumor-associated marker which represents a good target for the design of anti-tumor vaccines. Several studies have established that the binding of some anti-Tn antibodies could be affected by the density of Tn determinant or/and by the amino acid residues neighboring O-glycosylation sites. In the present study, using synthetic Tn-based vaccines, we have generated a panel of anti-Tn monoclonal antibodies. Analysis of their binding to various synthetic glycopeptides, modifying the amino acid carrier of the GalNAc(*) (Ser* vs Thr*), showed subtle differences in their fine specificities. We found that the recognition of these glycopeptides by some of these MAbs was strongly affected by the Tn backbone, such as a S*S*S* specific MAb (15G9) which failed to recognize a S*T*T* or a T*T*T* structure. Different binding patterns of these antibodies were also observed in FACS and Western blot analysis using three human cancer cell lines (MCF-7, LS174T and Jurkat). Importantly, an immunohistochemical analysis of human tumors (72 breast cancer and 44 colon cancer) showed the existence of different recognition profiles among the five antibodies evaluated, demonstrating that the aglyconic part of the Tn structure (Ser vs Thr) plays a key role in the anti-Tn specificity for breast and colon cancer detection. This new structural feature of the Tn antigen could be of important clinical value, notably due to the increasing interest of this antigen in anticancer vaccine design as well as for the development of anti-Tn antibodies for in vivo diagnostic and therapeutic strategies.     

Molecular characterization of totiviruses in Xanthophyllomyces dendrorhous

ABSTRACT: BACKGROUND: Occurrence of extrachromosomal dsRNA elements has been described in the red-yeast Xanthophyllomyces dendrorhous, with numbers and sizes that are highly variable among strains with different geographical origin. The studies concerning to the encapsidation of viral-like particles and dsRNA-curing have suggested that some dsRNAs are helper viruses, while others are satellite viruses. However, the nucleotide sequences and functions of these dsRNA are still unknown. In this work, the nucleotide sequences of four dsRNAs of the strain UCD 67-385 of X. dendrorhous were determined, and their identities and genome structures are proposed. Based on this molecular data, the dsRNAs of different strains of X. dendrorhous were analyzed. RESULTS: The complete sequences of L1, L2, S1 and S2 dsRNAs of X. dendrorhous UCD 67-385 were determined, finding two sequences for L1 dsRNA (L1A and L1B). Several ORFs were uncovered in both S1 and S2 dsRNAs, but no homologies were found for any of them when compared to the database. Instead, two ORFs were identified in each L1A, L1B and L2 dsRNAs, whose deduced amino acid sequences were homologous with a major capsid protein (5'-ORF) and a RNA-dependent RNA polymerase (3'-ORF) belonging to the Totivirus family. The genome structures of these dsRNAs are characteristic of Totiviruses, with two overlapped ORFs (the 3'-ORF in the -1 frame with respect to the 5'-ORF), with a slippery site and a pseudoknot in the overlapped regions. These structures are essential for the synthesis of the viral polymerase as a fusion protein with the viral capsid protein through -1 ribosomal frameshifting. In the RNase protection analysis, all the dsRNAs in the four analyzed X. dendrorhous strains were protected from enzymatic digestion. The RT-PCR analysis revealed that, similar to strain UCD 67-385, the L1A and L1B dsRNAs coexist in the strains VKM Y-2059, UCD 67-202 and VKM Y-2786. Furthermore, determinations of the relative amounts of L1 dsRNAs using two-step RT-qPCR revealed a 40-fold increment of the ratio L1A/L1B in the S2 dsRNA-cured strain compared to its parental strain. CONCLUSIONS: Three totiviruses, named as XdV-L1A, XdV-L1B and XdV-L2, were identified in the strain UCD 67-385 of X. dendrorhous. The viruses XdV-L1A and XdV-L1B were also found in other three X. dendrorhous strains. Our results suggest that the smaller dsRNAs (named XdRm-S1 and XdRm-S2) of strain UCD 67-385 are satellite viruses, and particularly that XdRm-S2 is a satellite of XdV-L1A.     

Cloning of complete genes for novel hydrolytic enzymes from Antarctic sea water bacteria by use of an improved genome walking technique

The increased demand for enzymes with new properties makes indispensable the development of easy and rapid strategies to obtain complete genes of new enzymes. Here a strategy is described which includes screening by PCR of new subtilases mediated by Consensus-Degenerate Hybrid Oligonucleotide Primers (CODEHOP) and an improved genome walking method to obtain the complete sequence of the identified genes. Existing methods of genome walking have many limitations, which make them inefficient and time consuming. We have developed an improved genome walking method with novel advances to get a simple, rapid and more efficient procedure based on cassette-ligation. Improvements consist basically in the possibility of a genomic DNA digestion with any restriction enzyme, blunting and 3' adenylation of digested DNA by Taq DNA polymerase to avoid self-circularization, followed by TA ligation of the adenine 3' overhanging end to the same unphosphorylated oligo-cassette. The efficiency of the genome walking method was demonstrated by finding the unknown ends of all gene fragments tested, previously obtained by CODEHOP-mediated PCR, including three subtilases (P4, P6 and P7), one xylanase and one lipase, from different strains of Antarctic marine bacteria.