RlmQ: a newly discovered rRNA modification enzyme bridging RNA modification and virulence traits in Staphylococcus aureus.

rRNA modifications play crucial roles in fine-tuning the delicate balance between translation speed and accuracy, yet the underlying mechanisms remain elusive. Comparative analyses of the rRNA modifications in taxonomically distant bacteria could help define their general, as well as species-specific, roles. In this study, we identified a new methyltransferase, RlmQ, in Staphylococcus aureus responsible for the Gram-positive specific m7G2601, which is not modified in Escherichia coli (G2574). We also demonstrate the absence of methylation on C1989, equivalent to E. coli C1962, which is methylated at position 5 by the Gram-negative specific RlmI methyltransferase, a paralog of RlmQ. Both modifications (S. aureus m7G2601 and E. coli m5C1962) are situated within the same tRNA accommodation corridor, hinting at a potential shared function in translation. Inactivation of S. aureus rlmQ causes the loss of methylation at G2601 and significantly impacts growth, cytotoxicity, and biofilm formation. These findings unravel the intricate connections between rRNA modifications, translation, and virulence in pathogenic Gram-positive bacteria.

Discovery of two distinct aminoacyl-tRNA synthetase complexes anchored to the Plasmodium surface tRNA import protein.

Aminoacyl-tRNA synthetases (aaRSs) attach amino acids to their cognate transfer RNAs. In eukaryotes, a subset of cytosolic aaRSs is organized into a multisynthetase complex (MSC), along with specialized scaffolding proteins referred to as aaRS-interacting multifunctional proteins (AIMPs). In Plasmodium, the causative agent of malaria, the tRNA import protein (tRip), is a membrane protein that participates in tRNA trafficking; we show that tRip also functions as an AIMP. We identified three aaRSs, the glutamyl-tRNA synthetase (ERS), glutaminyl-tRNA synthetase (QRS), and methionyl-tRNA synthetase (MRS), which were specifically coimmunoprecipitated with tRip in Plasmodium berghei blood stage parasites. All four proteins contain an N-terminal glutathione-S-transferase (GST)-like domain that was demonstrated to be involved in MSC assembly. In contrast to previous studies, further dissection of GST-like interactions identified two exclusive heterotrimeric complexes: the Q-complex (tRip-ERS-QRS) and the M-complex (tRip-ERS-MRS). Gel filtration and light scattering suggest a 2:2:2 stoichiometry for both complexes but with distinct biophysical properties and mutational analysis further revealed that the GST-like domains of QRS and MRS use different strategies to bind ERS. Taken together, our results demonstrate that neither the singular homodimerization of tRip nor its localization in the parasite plasma membrane prevents the formation of MSCs in Plasmodium. Besides, the extracellular localization of the tRNA-binding module of tRip is compensated by the presence of additional tRNA-binding modules fused to MRS and QRS, providing each MSC with two spatially distinct functions: aminoacylation of intraparasitic tRNAs and binding of extracellular tRNAs. This unique host-pathogen interaction is discussed.

Variable Intrinsic Expression of Immunoregulatory Biomarkers in Breast Cancer Cell Lines, Mammospheres, and Co-Cultures.

Advances in immunotherapy have increased interest in knowing the role of the immune system in breast cancer (BC) pathogenesis. Therefore, immune checkpoints (IC) and other pathways related to immune regulation, such as JAK2 and FoXO1, have emerged as potential targets for BC treatment. However, their intrinsic gene expression in vitro has not been extensively studied in this neoplasia. Thus, we evaluated the mRNA expression of tumor-cell-intrinsic CTLA-4, PDCD1 (PD1), CD274 (PD-L1), PDCD1LG2 (PD-L2), CD276 (B7-H3), JAK2, and FoXO1 in different BC cell lines, derived mammospheres, and co-cultures with peripheral blood mononuclear cells (PBMCs) by real-time quantitative polymerase chain reaction (qRT-PCR). Our results showed that intrinsic CTLA-4, CD274 (PD-L1), and PDCD1LG2 (PD-L2) were highly expressed in triple-negative cell lines, while CD276 was predominantly overexpressed in luminal cell lines. In contrast, JAK2 and FoXO1 were under-expressed. Moreover, high levels of CTLA-4, PDCD1 (PD1), CD274 (PD-L1), PDCD1LG2 (PD-L2), and JAK2 were found after mammosphere formation. Finally, the interaction between BC cell lines and peripheral blood mononuclear cells (PBMCs) stimulates the intrinsic expression of CTLA-4, PCDC1 (PD1), CD274 (PD-L1), and PDCD1LG2 (PD-L2). In conclusion, the intrinsic expression of immunoregulatory genes seems very dynamic, depending on BC phenotype, culture conditions, and tumor-immune cell interactions.

Tec1, a member of the TEA transcription factors family, is involved in virulence and basidiocarp development in Ustilago maydis.

The life cycle of Ustilago maydis involves alternation of a haploid saprophytic yeast-like stage and a dikaryotic hyphal virulent form. Under in vitro conditions, basidiocarps are formed. Analysis of the transcriptional network of basidiocarp formation revealed the possible involvement of a Tec transcription factor (Tec1, UMAG_02835) in the process. In some Ascomycota, Tec factors are involved in mycelial formation, pathogenesis, and interaction with other regulatory elements, but their role in Basidiomycota species is almost unknown. Accordingly, we proceeded to determine the role of this gene in U. maydis by its mutation. Tec1 was found to be a crucial factor for normal mating, basidiocarp development, and virulence, all of the functions related to the dikaryotic stage dependent of the b genes, whereas dimorphism and resistance to different stress conditions occurring in the haploid stage were not affected in tec1 mutants. The observation that mutants showed a low residual wild-type phenotype suggests the presence of a secondary mechanism that partially compensates the loss of Tec1.

Solanum tuberosum Microtuber Development under Darkness Unveiled through RNAseq Transcriptomic Analysis.

Potato microtuber (MT) development through in vitro techniques are ideal propagules for producing high quality potato plants. MT formation is influenced by several factors, i.e., photoperiod, sucrose, hormones, and osmotic stress. We have previously developed a protocol of MT induction in medium with sucrose (8% w/v), gelrite (6g/L), and 2iP as cytokinin under darkness. To understand the molecular mechanisms involved, we performed a transcriptome-wide analysis. Here we show that 1715 up- and 1624 down-regulated genes were involved in this biological process. Through the protein-protein interaction (PPI) network analyses performed in the STRING database (v11.5), we found 299 genes tightly associated in 14 clusters. Two major clusters of up-regulated proteins fundamental for life growth and development were found: 29 ribosomal proteins (RPs) interacting with 6 PEBP family members and 117 cell cycle (CC) proteins. The PPI network of up-regulated transcription factors (TFs) revealed that at least six TFs-MYB43, TSF, bZIP27, bZIP43, HAT4 and WOX9-may be involved during MTs development. The PPI network of down-regulated genes revealed a cluster of 83 proteins involved in light and photosynthesis, 110 in response to hormone, 74 in hormone mediate signaling pathway and 22 related to aging.

Analysis of the photoreceptors involved in the light-depending basidiocarp formation in Ustilago maydis.

We have described that formation of basidiocarps by Ustilago maydis requires illumination. In the current research, we have proceeded to analyze what kind of light receptors are involved in this phenomenon. Accordingly, we investigated whether the homologues of the White Collar (WC), and the phytochrome (PHY) genes played a role in this process. Mutants deficient in either one of the three U. maydis WC homologue genes (WCO1a, WCO1b, WCO2), or the phytochrome-encoding the PHY gene were obtained. Phenotypic analysis of the mutants showed that ∆wco1a mutants formed similar numbers of basidiocarps than wild-type strain, whereas ∆wco1b mutants were severely affected in basidiocarp formation when illuminated with white, blue or red light. ∆wco2 and ∆phy1 mutants did not form basidiocarps under any illumination condition. These data indicate that Wco1a is the main blue light receptor, and Wco1b may operate as a secondary blue light receptor; Phy1 is the red light receptor, and Wco2 the transcription factor that controls the photo stimulation of the genes involved in the formation of fruiting bodies. It is suggested that effectiveness of the light receptors depends on the whole structure of the complex, possibly, because their association is necessary to maintain their functional structure.

Agrobacterium rhizogenes-mediated transformation of grain (Amaranthus hypochondriacus) and leafy (A. hybridus) amaranths.

KEY MESSAGE: Transgenic A. hypochondriacus and A. hybridus roots were generated. Further, a distinct plant regeneration program via somatic embryos produced from hairy roots was established. Work was implemented to develop an optimized protocol for root genetic transformation of the three grain amaranth species and A. hybridus, their presumed ancestor. Transformation efficiency was species-specific, being higher in A. hypochondriacus and followed by A. hybridus. Amaranthus cruentus and A. caudatus remained recalcitrant. A reliable and efficient Agrobacteruim rhizogenes-mediated transformation of these species was established using cotyledon explants infected with the previously untested BVG strain. Optimal OD600 bacterial cell densities were 0.4 and 0.8 for A. hypochondriacus and A. hybridus, respectively. Hairy roots of both amaranth species were validated by the amplification of appropriate marker genes and, when pertinent, by monitoring green fluorescent protein emission or ß-glucuronidase activity. Embryogenic calli were generated from A. hypochondriacus rhizoclones. Subsequent somatic embryo maturation and germination required the activation of cytokinin signaling, osmotic stress, red light, and calcium incorporation. A crucial step to ensure the differentiation of germinating somatic embryos into plantlets was their individualization and subcultivation in 5/5 media containing 5% sucrose, 5 g/L gelrite, and 0.2 mg/L 2-isopentenyladenine (2iP) previously acidified to pH 4.0 with phosphoric acid, followed by their transfer to 5/5 + 2iP media supplemented with 100 mg/L CaCl2. These steps were strictly red light dependent. This process represents a viable protocol for plant regeneration via somatic embryo germination from grain amaranth transgenic hairy roots. Its capacity to overcome the recalcitrance to genetic transformation characteristic of grain amaranth has the potential to significantly advance the knowledge of several unresolved biological aspects of grain amaranths.

Patients' quality of life: Validation of the functional assessment of cancer therapy-bone marrow transplant (FACT-BMT) instrument for the Mexican population.

OBJECTIVE: The functional assessment of cancer therapy-bone marrow transplant (FACT-BMT) is a widely used instrument to assess quality of life (QOL) in hematopoietic stem cell transplant (HSCT) patients, but there is little evidence of its validity in Latin American populations. This study evaluated the psychometric properties of the Spanish language version of the FACT-BMT in Mexican patients. METHOD: First, the original version was piloted with 15 HSCT patients to obtain an adequate cultural version, resulting in the adaptation of one item. After that, the new version was completed by 139 HSCT patients. RESULTS: The results showed a FACT factor structure that explains 70.84% of the total variance, a factor structure similar to the original FACT structure, and with a high internal consistency (Cronbach's alpha = 0.867). For the BMT subscale, the best factor structure included 17 items which explain 61.65% of the total variance with an adequate internal consistency (Cronbach's alpha = 0.696). SIGNIFICANCE OF THE RESULTS: The FACT-BMT was found to be a valid and reliable instrument to evaluate QOL in Mexican patients. Our results constitute new FACT-BMT empirical evidence that supports its clinical and research uses.

Validation of an instrument to assess health care quality (FACIT-TS-PS) in cancer patients.

INTRODUCTION: Cancer patient satisfaction with the healthcare team is of great relevance for assessing the quality of the care provided by the health system. In Mexico, no valid and reliable tool is available to assess this construct. OBJECTIVE: To validate the Functional Assessment of Chronic Illness Therapy-Treatment Satisfaction-Patient Satisfaction (FACIT-TS-PS) instrument, version 4, in cancer patients. METHOD: Cross-sectional design, non-probability convenience sampling. The sample consisted of 200 cancer-diagnosed patients, with mean age of 45.86 ± 15.01 years. Exploratory and confirmatory factor analyses were conducted. RESULTS: The exploratory factor analysis identified four factors, with a Cronbach alpha of 0.945, and an explained variance of 68.15 %. The confirmatory factor analysis indicated that the proposed theoretical model adjusts to the data with an error close to zero and, in addition, it is balanced and carefully measures overall patient satisfaction with the treatment. CONCLUSION: FACIT-TS-PS was shown to be a valid and reliable instrument for use in clinical care and research in Mexican cancer patients. Its use is recommended in the evaluation of oncology multidisciplinary healthcare teams in Mexico.

INTRODUCCIÓN: La satisfacción del paciente oncológico con el equipo de salud es de relevancia para evaluar la calidad de la atención del sistema de salud. En México no se dispone de una herramienta válida y confiable para evaluar este constructo. OBJETIVO: Validar el Instrumento de Evaluación de Funcionalidad en el Tratamiento para Enfermedades Crónicas-Satisfacción con el Tratamiento-Satisfacción del Paciente (FACIT-TS-PS) versión 4, en pacientes mexicanos con cáncer. MÉTODO: Diseño transversal, muestreo no probabilístico, por disponibilidad. La muestra consistió en 200 pacientes diagnosticados con cáncer, con edad promedio de 45.86 ± 15.01 años. Se realizó un análisis factorial exploratorio y confirmatorio. RESULTADOS: Se identificaron cuatro factores con un alfa de Cronbach de 0.945 y una varianza explicada de 68.15 %. El análisis factorial confirmatorio indicó que el modelo teórico propuesto se ajusta a los datos con error próximo a cero y que, además, es equilibrado y mide cuidadosamente la satisfacción global del paciente con el tratamiento. CONCLUSIÓN: FACIT-TS-PS mostró ser un instrumento válido y confiable para su uso en la atención clínica e investigación dirigida a pacientes mexicanos con cáncer. Se recomienda su utilización en la evaluación de equipos de salud multidisciplinarios en oncología en México.

In vitro treatment of Enterococcus faecalis with calcium hydroxide impairs phagocytosis by human macrophages.

OBJECTIVE: Monocyte-derived macrophages (MDMs) ability to phagocytize and produce nitric oxide (NO) was tested against root-canal strains of Enterococcus faecalis submitted to alkaline stress. Root-canal strains were also compared with urine Enterococci. MATERIALS AND METHODS: Enterococcus faecalis were stressed with alkaline-BHI broth and incubated in vitro at a cell/bacteria ratio of 1:5. Phagocytosis was analyzed by fluorescence microscopy using acridine orange stain, and NO concentration was measured in supernatants. RESULTS AND CONCLUSIONS: Alkaline-stress significantly impaired MDMs phagocytosis of E. faecalis strains analyzed, except in ATCC4083 isolated from a pulpless tooth, but NO production was unchanged. Comparison of different strains showed the urine isolate had higher NO levels than root canal strains. Alterations in the bacterial cell wall structures after alkaline-stress possibly made bacteria less recognizable and phagocytized by MDMs but did not affect their ability to activate NO production. Furthermore, root canal strains elicited different responses by immune cells compared with strains from urine. Clinically, impaired phagocytosis of E. faecalis could contribute to their persistence in root canal systems previously treated with calcium hydroxide.

Development of an Efficient Protocol to Obtain Transgenic Coffee, Coffea arabica L., Expressing the Cry10Aa Toxin of Bacillus thuringiensis.

This report presents an efficient protocol of the stable genetic transformation of coffee plants expressing the Cry10Aa protein of Bacillus thuringiensis. Embryogenic cell lines with a high potential of propagation, somatic embryo maturation, and germination were used. Gene expression analysis of cytokinin signaling, homedomains, auxin responsive factor, and the master regulators of somatic embryogenesis genes involved in somatic embryo maturation were evaluated. Plasmid pMDC85 containing the cry10Aa gene was introduced into a Typica cultivar of C. arabica L. by biobalistic transformation. Transformation efficiency of 16.7% was achieved, according to the number of embryogenic aggregates and transgenic lines developed. Stable transformation was proven by hygromycin-resistant embryogenic lines, green fluorescent protein (GFP) expression, quantitative analyses of Cry10Aa by mass spectrometry, Western blot, ELISA, and Southern blot analyses. Cry10Aa showed variable expression levels in somatic embryos and the leaf tissue of transgenic plants, ranging from 76% to 90% of coverage of the protein by mass spectrometry and from 3.25 to 13.88 µg/g fresh tissue, with ELISA. qPCR-based 2-ΔΔCt trials revealed high transcription levels of cry10Aa in somatic embryos and leaf tissue. This is the first report about the stable transformation and expression of the Cry10Aa protein in coffee plants with the potential for controlling the coffee berry borer.

CD44 and ALDH1 immunoexpression as prognostic indicators of invasion and metastasis in oral squamous cell carcinoma.

BACKGROUND: Tumour metastasis has been associated with cancer stem cells, a small population with stem-like cells properties, higher rate of migration and metastatic potential compared to cells from the tumour bulk. Our aim was to evaluate the immunoexpression of the putative cancer stem cell biomarkers ALDH1 and CD44 in primary tumour and corresponding metastatic lymph nodes. METHODS: Tumour tissue specimens (n = 50) and corresponding metastatic lymph nodes (n = 25) were surgically obtained from 50 patients with oral squamous cell carcinoma and submitted to immunohistochemistry. CD44 and ALDH1 were semi-quantitatively scored according to the proportion and intensity of positive cells within the invasive front and metastatic lymph nodes as a whole. A combined score was obtained by multiplying both parameters and later dichotomized into a final score classified as low (≤2) or high (>2) immunoexpression. RESULTS: ALDH1 immunoexpression and CD44 immunoexpression were detected in both tumour sites, although the means of ALDH1 (P = .0985) and CD44 (P = .4220) cells were higher in metastasis compared to primary tumours. ALDH1high was positively associated (P = .0184) with angiolymphatic invasion, while CD44high was positively associated (P = .0181) with metastasis (N+). At multivariate analysis, CD44 significantly increased the odds of lymph node metastasis, regardless of T stage (OR = 8.24; 1.64-65.64, P = .0088). CONCLUSIONS: CD44 immunoexpression was a significant predictor of lymph node metastasis, while ALDH1high immunostaining was associated with angiolymphatic invasion. Altogether, it suggests that immunoexpression of CD44 and ALDH1 links the cancer stem cell phenotype with oral squamous cell carcinoma invasion and metastasis.

Epidemiological Data on the Nutritional Status of Cancer Patients Receiving Treatment with Concomitant Chemoradiotherapy, Radiotherapy or Sequential Chemoradiotherapy to the Abdominopelvic Area.

Cancer patients are particularly susceptible to undernourishment so associated weight loss is frequent. Approximately 15% of patients lose >10% of their usual body weight, 40-80% become undernourished, and about 20% die as a result. Well-nourished patients have a higher survival rate when compared with patients at risk of undernourishment (19.9 vs. 3.7 months); hence, nutritional intervention is pivotal. Undernourishment negatively influences the patient's prognosis, and its prevalence depends on the tumor type and location, disease stage, treatment, and the applied nutritional evaluation tool. During abdominopelvic radiotherapy, up to 90% of patients experience symptoms of varying severity; weight loss during radiotherapy is an early indicator of nutritional deterioration, and he the use of radiation is associated with a higher likelihood of undernourishment. In patients with gynecological malignancies, 12.5-54% are malnourished before receiving oncological treatment, worsening after treatment in 35.8-82% of cases. There is also deterioration of the nutritional status in patients with colorectal cancer once pelvic radiotherapy is initiated, whereby 50% of cases are malnourished at the beginning of treatment, and 66.7% are so when it ends. Although there are notable differences in the impact of radiotherapy on weight according to the radiated region, 88% patients receiving abdominal radiotherapy were found to lose weight compared to 38% of patients whose treatment was limited to the pelvis.

Advanced Squamous Cell Carcinoma of the Head and Neck: The Current Role of Cetuximab.

We review clinical trials of squamous cell carcinoma of the head and neck (SCCHN) to address the current and potential uses of cetuximab (CTX). PubMed was reviewed to identify papers published between 2010 and 2016. The search terms used were "cetuximab" and "head and neck cancer." A total of 634 articles were identified. Phase II or III studies with CTX in patients with advanced SCCHN without treatment or with recurrent/metastatic tumors were selected. Forty-six registries were obtained. Information was critically reviewed and relevant information presented. As definitive treatment of advanced squamous cells carcinomas and as palliative treatment of recurrent/metastatic disease, CTX alone or associated with chemotherapy and/or radiotherapy is an alternative to chemoradiotherapy because of its distinct and favorable toxicity profile.

Zoledronic acid in the treatment of metastatic breast cancer.

Bone is the most common location of metastatic disease. Approximately 80% of all bone metastases are observed in patients with breast or prostate tumours and are responsible for more than 300 000 deaths every year. Treatment of malignant bone disease with bisphosphonates has been shown to reduce bone events and delay their onset, and several reviews and meta-analyses have confirmed the benefit of these drugs in controlling bone metastases. Zoledronic acid is a bisphosphonate that has been shown to delay or prevent the development of skeletal-related events in patients with bone metastases. Furthermore, compared with other bisphosphonates, zoledronic acid has also shown better pain control and various studies also suggest an improvement in quality of life, although with no impact on overall survival. The duration and optimal regimen for long-term zoledronic acid therapy have not yet been defined, but some studies suggest that continuing zoledronic acid therapy for more than 2 years could also extend its beneficial effect.

Pharyngeal mucosal closure in total laryngectomy: comparison between vertical and T-shaped closure.

Objective: The objective of this study was to compare the outcomes vertical and T-shaped pharyngoplasty closure techniques after total laryngectomy (TL) and to evaluate the factors associated with the development of pharyngocutaneous fistula. Materials and methods: We performed a retrospective study that included patients with a histopathological diagnosis of laryngeal cancer that underwent TL between 2009 and 2021. Results: Fifty-seven patients were included in the study. A total of 14 patients underwent a vertical closure of the neopharynx (24.6%), while 43 patients underwent a T-shaped closure (74.4%). Pharyngocutaneous fistula was the most common complication, observed in 40.4% of cases (n = 23). No difference in the rate of complications was observed between groups, with the exception of tracheal dehiscence which was reduced in patients with T-shaped closure (n = 2, 4.7% vs. n = 5, 35.7%, p = 0.002). Diabetes mellitus was more frequently observed in patients withthe development of pharyngocutaneous fistula (n = 7, 30.4% vs. n = 3, 8.8%, p = 0.03). Conclusions: Although complicationswere lower in the T-shaped closure group, we could not establish the superiority of either technique.

Objetivo: Evaluar los desenlaces de la técnica vertical en comparación con la técnica en T para el cierre de faringoplastia posterior a una laringectomía total, y evaluar los factores asociados con el desarrollo de fístula faringocutánea. Método: Estudio retrospectivo de pacientes con diagnóstico de cancer de laringe a quienes se realizó laringectomía total como tratamiento, de 2009 a 2021. Resultados: Se incluyeron 57 pacientes. A 14 (24.6%) se les realizó una faringoplastia con cierre en T y a 43 (74.4%) un cierre vertical. La fístula faringocutánea fue la complicación más frecuente, presente en el 40.4% de los casos (n = 23). No se observaron diferencias en el desarrollo de complicaciones entre grupos, con excepción de la dehiscencia traqueal, la cual fue menos frecuente en el grupo de cierre en T (n = 2, 4.7% vs. n = 5, 35.7%; p = 0.002). La diabetes mellitus se asoció con el desarrollo de fístula faringocutánea (n = 7, 30.4% vs. n = 3, 8.8%; p = 0.03). Conclusiones: Aunque se observó una tendencia a una disminución de las complicaciones en el grupo de cierre en T, no se encontró superioridad de una técnica sobre otra.

Ribosome Profiling Methods Adapted to the Study of RNA-Dependent Translation Regulation in Staphylococcus aureus.

Noncoding RNAs, including regulatory RNAs (sRNAs), are instrumental in regulating gene expression in pathogenic bacteria, allowing them to adapt to various stresses encountered in their host environments. Staphylococcus aureus is a well-studied model for RNA-mediated regulation of virulence and pathogenicity, with sRNAs playing significant roles in shaping S. aureus interactions with human and animal hosts. By modulating the translation and/or stability of target mRNAs, sRNAs regulate the synthesis of virulence factors and regulatory proteins required for pathogenesis. Moreover, perturbation of the levels of RNA modifications in two other classes of noncoding RNAs, rRNAs, and tRNAs, has been proposed to contribute to stress adaptation. However, the study of how these various factors affect translation regulation has often been restricted to specific genes, using in vivo reporters and/or in vitro translation systems. Genome-wide sequencing approaches offer novel perspectives for studying RNA-dependent regulation. In particular, ribosome profiling methods provide a powerful resource for characterizing the overall landscape of translational regulation, contributing to a better understanding of S. aureus physiopathology. Here, we describe protocols that we have adapted to perform ribosome profiling in S. aureus.

Hypertension as predictive factor for bevacizumab-containing first-line therapy in metastatic breast and colorectal cancer in BRECOL (GEICAM/2011-04) study.

BACKGROUND: Retrospective data suggest an association between bevacizumab efficacy and the incidence of arterial hypertension (AHT). Additionally, epigenetic mechanisms have been related to AHT. METHODS: This prospective observational study conducted by GEICAM Spanish Breast Cancer Research Group included metastatic breast (MBC) or colorectal (mCRC) cancer patients treated with bevacizumab-containing chemotherapy as first-line treatment. Blood pressure (BP) levels were measured (conventional and 24-h Holter monitoring) at baseline and up to cycle 3. Primary endpoint assessed BP levels increase as predictive factor for progression-free survival (PFS). Germline DNA methylation profile was explored in pre-treatment blood samples; principal component analysis was used to define an epigenetic predictive score for increased BP levels. RESULTS: From Oct-2012 to Jul-2016, 143 (78 MBC and 65 mCRC) patients were included. The incidence of AHT according to guidelines was neither predictive of PFS nor of best overall tumor response (BOR). No statistically significant association was observed with systolic BP nor diastolic BP increment for PFS or BOR. Grade 3 and 4 adverse events were observed in 37 and 5% of patients, respectively. We identified 27 sites which baseline methylation status was significantly associated to BP levels increase secondary to bevacizumab-containing chemotherapy. CONCLUSIONS: Neither the frequency of AHT nor the increase of BP levels were predictive of efficacy in MBC and mCRC patients treated with bevacizumab-containing chemotherapy. CLINICAL TRIAL REGISTRY: ClinicalTrials.gov Identifier: NCT01733628.

Editing of SlWRKY29 by CRISPR-activation promotes somatic embryogenesis in Solanum lycopersicum cv. Micro-Tom.

At present, the development of plants with improved traits like superior quality, high yield, or stress resistance, are highly desirable in agriculture. Accelerated crop improvement, however, must capitalize on revolutionary new plant breeding technologies, like genetically modified and gene-edited crops, to heighten food crop traits. Genome editing still faces ineffective methods for the transformation and regeneration of different plant species and must surpass the genotype dependency of the transformation process. Tomato is considered an alternative plant model system to rice and Arabidopsis, and a model organism for fleshy-fruited plants. Furthermore, tomato cultivars like Micro-Tom are excellent models for tomato research due to its short life cycle, small size, and capacity to grow at high density. Therefore, we developed an indirect somatic embryo protocol from cotyledonary tomato explants and used this to generate epigenetically edited tomato plants for the SlWRKY29 gene via CRISPR-activation (CRISPRa). We found that epigenetic reprogramming for SlWRKY29 establishes a transcriptionally permissive chromatin state, as determined by an enrichment of the H3K4me3 mark. A whole transcriptome analysis of CRISPRa-edited pro-embryogenic masses and mature somatic embryos allowed us to characterize the mechanism driving somatic embryo induction in the edited tomato cv. Micro-Tom. Furthermore, we show that enhanced embryo induction and maturation are influenced by the transcriptional effector employed during CRISPRa, as well as by the medium composition and in vitro environmental conditions such as osmotic components, plant growth regulators, and light intensity.