Case Report: Characterizing the Role of the STXBP2-R190C Monoallelic Mutation Found in a Patient With Hemophagocytic Syndrome and Langerhans Cell Histiocytosis

Histiocitosi de cèl·lules de Langerhans; Desgranulació; Exocitosi de grànuls líticsHistiocitosis de células de Langerhans; Desgranulación; Exocitosis de gránulos líticosLangerhans-cell-histiocytosis; Degranulation; Lytic granule exocytosisHemophagocytic lymphohistiocytosis (HLH) is a life-threatening hyperinflammatory disorder. HLH can be considered as a threshold disease depending on the trigger and the residual NK-cell cytotoxicity. In this study, we analyzed the molecular and functional impact of a novel monoallelic mutation found in a patient with two episodes of HLH. A 9-month-old child was diagnosed at 2 months of age with cutaneous Langerhans cell histiocytosis (LCH). After successful treatment, the patient developed an HLH episode. At 16 month of age, the patient went through an HSCT losing the engraftment 5 months later concomitant with an HLH relapse. The genetic study revealed a monoallelic mutation in the STXBP2 gene (.pArg190Cys). We transfected COS7 cells to analyze the STXBP2-R190C expression and to test the interaction with STX11. We used the RBL-2H3 cell line expressing STXBP2-WT-EGFP or R190C-EGFP for degranulation assays. Mutation STXBP2-R190C did not affect protein expression or interaction with syntaxin-11. However, we have demonstrated that STXBP2-R190C mutation diminishes degranulation in the RBL-2H3 cell line compared with the RBL-2H3 cell line transfected with STXBP2-WT or nontransfected. These results suggest that STXBP2-R190C mutation acts as a modifier of the degranulation process producing a decrease in degranulation. Therefore, under homeostatic conditions, the presence of one copy of STXBP2-R190 could generate sufficient degranulation capacity. However, it is likely that early in life when adaptive immune system functions are not sufficiently developed, an infection may not be resolved with this genetic background, leading to a hyperinflammation syndrome and eventually develop HLH. This analysis highlights the need for functional testing of new mutations to validate their role in genetic susceptibility and to establish the best possible treatment for these patients.This work was funded by the Instituto de Salud Carlos III, grants PI17/00660 and PI18/00346, co-financed by the European Regional Development Fund (ERDF)

Haploidentical vs. HLA-matched donor hematopoietic stem-cell transplantation for pediatric patients with acute lymphoblastic leukemia in second remission: A collaborative retrospective study of the Spanish Group for Bone Marrow Transplantation in Children (GETMON/GETH) and the Spanish Childhood Relapsed ALL Board (ReALLNet)

IntroductionStudies addressing the role of haploidentical as alternative to HLA-matched donors for stem cell transplantation (SCT) often include patients with diverse hematological malignancies in different remission statuses.MethodsWe compared outcomes of children with acute lymphoblastic leukemia (ALL) undergoing SCT in second complete remission (CR2) from haploidentical (n = 25) versus HLA-matched donor (n = 51).ResultsPatients were equally distributed across both groups according to age, immunophenotype, time to and site of relapse, relapse risk-group allocation, and minimal residual disease (MRD) before SCT. Incidence of graft failure, acute graft versus host disease (GVHD), and other early complications did not differ between both groups. We found no differences in overall survival (58.7% versus 59.5%; p = .8), leukemia free survival (LFS) (48% versus 36.4%; p = .5), event free survival (40% versus 34.4%; p = .69), cumulative incidence (CI) of subsequent relapse (28% versus 40.9%; p = .69), treatment related mortality (24% versus 23.6%; p = .83), CI of cGVHD (4.5% versus 18.7%; p = .2), and chronic GVHD-free and leukemia-free survival (44% versus 26.3%; p = .3) after haploidentical donor SCT. Chronic GVHD (HR = 0.09; p=.02) had protective impact, and MRD ≥ 0.01% before SCT (HR = 2.59; p=.01) had unfavorable impact on LFS.DiscussionThese results support the role of haploidentical donor SCT in children with ALL in CR2

Definición del núcleo optimizado de la colección de conservación del manzano español

5 Pags.- 1 Tabl.Este trabajo tiene como objetivo determinar la estrategia más adecuada para la selección del conjunto mínimo de accesiones (núcleo optimizado) que represente eficientemente la variación genética del manzano conservado en las colecciones españolas. Este núcleo optimizado mediante criterios genéticos constituirá la base de la colección nuclear de conservación, que podrá ser complementado con accesiones seleccionadas por otros criterios (morfo-fisiológicos, agronómicos, valor histórico, etc.). Se ha evaluado la eficiencia de estrategias de selección por búsqueda local estocástica avanzada (ASLS) que diferían tanto por el tamaño final del núcleo como por la combinación (y peso relativo) de las medidas de distancia genética y riqueza alélica a optimizar. Las estrategias empleadas han proporcionado núcleos optimizados con grandes diferencias en la diversidad conservada, así como en el nivel de representación de la estructura genética general. Teniendo en cuenta el uso principal de la colección nuclear, la estrategia que ofrece un mejor equilibrio entre representatividad y adecuación al uso es la que combina la optimización de la distancia media entre cada accesión de la colección y la entrada en el núcleo más cercana con el índice de Shannon y la recuperación de alelos.Este trabajo ha sido financiado por los proyectos INIA RF2011-00017-C05-00 y RTA2015-00052-C02-00Peer reviewe

Development of a standardized methodology for phenotypical characterizations in apple

4 Pags.- 2 Tabls. Articles derived from XIV EUCARPIA Symposium on Fruit Breeding and Genetics (Bologna,Italy. June 14-18 2015) . The definitive version is available at: http://www.actahort.org/index.htmThe description of phenotypic traits in apple cultivars is generally performed using internationally agreed descriptors such as UPOV guidelines, which defines for each trait several states of expression. However, it is not always possible to classify a cultivar unambiguously using those guidelines, because in practice the states are not clearly enough defined or the example cultivars are not always available in the collections. This work presents the results of a harmonization project performed by the teams responsible of the main apple germplasm collections in Spain. The objective was to develop a standardized method for the 57 traits included in the TG/14/9 UPOV guidelines for apple characterization, defining their states of expression in a clear and unambiguous way for Spanish germplasm. Phenotypic data collected for more than 1,600 accessions from Spanish collections were used and the method to define each state depended on the type of expression. For quantitative traits the number of states and their limits were defined according to the variability that exists within and between accessions. For qualitative traits, high-resolution images clearly depicting each state were selected. A standardized characterization protocol for the 57 traits of apple germplasm has been provided, enabling to comparing properly the phenotypes of Spanish genetic resources.This Project has been funded by the Spanish Ministry of Science and Innovation/National Institute for Agricultural and Food Research and Technology (RF2011- 00017-C05-00).Peer reviewe

Development of a standardized methodology for phenotypical characterizations in apple

1 pag.This work presents the results of a harmonization project performed by the teams responsible of the main apple germplasm collections in Spain. The objective was to develop a standardized method for the 57 traits included in the TG/14/9 UPOV guidelines for apple characterization, defining their states of expression in a clear and unambiguous way for Spanish germplasm.Peer reviewe

Time to Switch to Second-line Antiretroviral Therapy in Children With Human Immunodeficiency Virus in Europe and Thailand.

Background: Data on durability of first-line antiretroviral therapy (ART) in children with human immunodeficiency virus (HIV) are limited. We assessed time to switch to second-line therapy in 16 European countries and Thailand. Methods: Children aged <18 years initiating combination ART (≥2 nucleoside reverse transcriptase inhibitors [NRTIs] plus nonnucleoside reverse transcriptase inhibitor [NNRTI] or boosted protease inhibitor [PI]) were included. Switch to second-line was defined as (i) change across drug class (PI to NNRTI or vice versa) or within PI class plus change of ≥1 NRTI; (ii) change from single to dual PI; or (iii) addition of a new drug class. Cumulative incidence of switch was calculated with death and loss to follow-up as competing risks. Results: Of 3668 children included, median age at ART initiation was 6.1 (interquartile range (IQR), 1.7-10.5) years. Initial regimens were 32% PI based, 34% nevirapine (NVP) based, and 33% efavirenz based. Median duration of follow-up was 5.4 (IQR, 2.9-8.3) years. Cumulative incidence of switch at 5 years was 21% (95% confidence interval, 20%-23%), with significant regional variations. Median time to switch was 30 (IQR, 16-58) months; two-thirds of switches were related to treatment failure. In multivariable analysis, older age, severe immunosuppression and higher viral load (VL) at ART start, and NVP-based initial regimens were associated with increased risk of switch. Conclusions: One in 5 children switched to a second-line regimen by 5 years of ART, with two-thirds failure related. Advanced HIV, older age, and NVP-based regimens were associated with increased risk of switch

Heat stress factors expressed during seed maturation differentially regulate seed longevity and seedling greening

Heat Stress Factor A9 (A9), a seed-specific transcription factor contributing to seed longevity, also enhances phytochrome-dependent seedling greening. The RNA-seq analyses of imbibed-seed transcripts here reported indicated potential additional effects of A9 on cryptochrome-mediated blue-light responses. These analyses also suggested that in contrast to the A9 effects on longevity, which require coactivation by additional factors as A4a, A9 alone might suffice for the enhancement of photomorphogenesis at the seedling stage. We found that upon its seed-specific overexpression, A9 indeed enhanced the expected blue-light responses. Comparative loss-of-function analyses of longevity and greening, performed by similar expression of dominant-negative and inactive forms of A9, not only confirmed the additional greening effects of A9, but also were consistent with A9 not requiring A4a (or additional factors) for the greening effects. Our results strongly indicate that A9 would differentially regulate seed longevity and photomorphogenesis at the seedling stage, A9 alone sufficing for both the phytochrome-and cryptochrome-dependent greening enhancement effects

Heat Stress Factors Expressed during Seed Maturation Differentially Regulate Seed Longevity and Seedling Greening

13 páginas.- 7 figuras.- 36 referencias.- Supplementary Materials: The following are available online at http://www.mdpi.com/2223-7747/9/3/335/s1.- This article belongs to the Special Issue Genetics of Seed Germination and GrowthHeat Stress Factor A9 (A9), a seed-specific transcription factor contributing to seed longevity, also enhances phytochrome-dependent seedling greening. The RNA-seq analyses of imbibed-seed transcripts here reported indicated potential additional effects of A9 on cryptochrome-mediated blue-light responses. These analyses also suggested that in contrast to the A9 effects on longevity, which require coactivation by additional factors as A4a, A9 alone might suffice for the enhancement of photomorphogenesis at the seedling stage. We found that upon its seed-specific overexpression, A9 indeed enhanced the expected blue-light responses. Comparative loss-of-function analyses of longevity and greening, performed by similar expression of dominant-negative and inactive forms of A9, not only confirmed the additional greening effects of A9, but also were consistent with A9 not requiring A4a (or additional factors) for the greening effects. Our results strongly indicate that A9 would differentially regulate seed longevity and photomorphogenesis at the seedling stage, A9 alone sufficing for both the phytochrome- and cryptochrome-dependent greening enhancement effects.This research was funded by FEDER (European Regional Development Fund) and by “Secretaría de Estado de Investigación, Desarrollo e Innovación. Ministerio de Economía, Industria y Competitividad” (Projects BIO2014-52303-R and BIO2017-82172-R

Characterization of chromatin accessibility during the seed to seedling transition in nicotiana tabacum

Poster presentado en el XVI Meeting of Plant Molecular Biology (RPMB) 14-16 sept. 2022, SevillaChromatin accessibility (CA) varies at different development stages, and despite regulation of the accessible genome being crucial for development in plants, related knowledge is scarce and mostly limited to Arabidopsis (Ding et al., 2022). Here, we globally profiled CA genome-wide in tobacco (Nicotiana tabacum), with the aim of analysing accessible regions in seeds immediately after germination. We performed formaldehyde-assisted isolation of regulatory elements coupled with high-throughput sequencing (FAIRE-seq; Simon et al., 2012). Sequencing was performed before exposure to light, which leads to seedling establishment following subsequent activation of photosynthetic autotrophic growth. We are also aiming to investigate the global effect on CA of HSFA9 (A9), a seed-specific transcription factor for which we also reported effects on selected gene loci at this stage, such as PHYA1, ATS1, and HY5 (see for example Prieto-Dapena et al., 2021, and the communication by Carranco et al. in this meeting). We performed preliminary low-coverage FAIRE-seq on seeds from two independent, transgenic, DS10:A9 lines that express A9 from a seed-specific promoter, together with the corresponding non-transgenic (NT) siblings. Sequencing reads were aligned to a chromosome-based reference genome derived from published data (Edwards et al., 2017). We consistently report enrichment of CA in the 5´-proximal regions of coding sequences (thus “promoters” with transcription start sites -TSS- and, close, cis-acting sequences). Indeed, the regions corresponding to putative promoters displayed ~4X more accessible sites (i.e., significant enrichment of CA) than expected by chance. We also observed accessible sites at more distal locations (around a third of our dataset was located more than 10Kb upstream of the closest gene). These might include trans-acting stage-specific enhancers involved in long-range interactions with relevant promoters, or point to unannotated genes. Following an ongoing deeper sequencing (c.a. 450 million reads), our data would reveal the genome-wide sequences involved in chromatin regulation during the seed to autotrophic-seedling transition, potentially unveiling differential chromatin-related effect(s) caused by A9 at this stage. Ding et al. (2022) DOI: 10.3389/fpls.2022.865361 Edwards et. al. (2017) DOI:10.1186/s12864-017-3791-6 Prieto-Dapena et al. (2021) Plant Biology 21 Worldwide Summit (Abstract ID: 1081784) Simon et al., (2012) DOI: 10.1038/nprot.2011.444This work has been funded by the European Regional Development Fund (FEDER) and the Spanish Secretariat of Research, Development and Innovation (BIO2017-82172-R and PID2020-112693RB-100/ AEI/10.13039/501100011033). Antonio Pérez-Cardona received a contract from Garantía Juvenil Andalucía 2021 (Ref. 37803).N