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1.
Sci Total Environ ; 914: 169653, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38176556

RESUMO

Rice is one of the major cereal crops in the world, contributing significantly towards the dietary energy and nutrition of more than half of the world's population. However, rice can also be a significant exposure route for inorganic arsenic (iAs). This risk is even greater if rice is cooked with iAs-contaminated water. Here, we quantified the effect of two cooking methods, excess water (EW) and parboiled and absorbed (PBA), on As species and essential nutrient elements (P, K, Mg, Fe, Zn, Mn, Cu, Se and Mo) in white, parboiled and brown rice cooked with As-safe (0.18 µg L-1) and As-spiked (10 and 50 µg L-1) tap water. Furthermore, we calculated the exposure risk using the margin of exposure (MOE) for both low (the UK) and high (Bangladesh) rice per capita consumption scenarios. The total micro and macronutrient content in cooked rice was measured using ICP-MS (Inductively Coupled Plasma Mass Spectrometry). An LC-ICP-MS (liquid chromatography-ICP-MS) method was used to quantify arsenic species. The results demonstrate that EW and PBA methods produced similar efficacy of iAs removal (54-58 %) for white and brown rice. However, the EW method was better at removing iAs from parboiled rice (∼50 %) than PBA (∼39 %). We found that cooked brown rice was superior to other rice types in many essential nutrient elements, and cooking methods significantly affected the loss of K, Fe, Cu and Mo. For both cooking methods, cooking with iAs-spiked water significantly increased iAs in all rice types: white > parboiled > brown. However, when using As-spiked water, the PBA method retained more iAs than EW. Our risk evaluations showed that cooking rice with 50 µg L-1 significantly raises the As-exposure of the Bangladesh population due to the high per capita rice consumption rate, reinforcing the importance of accessing As-safe water for cooking.


Assuntos
Arsênio , Oryza , Arsênio/análise , Oryza/química , Contaminação de Alimentos/análise , Culinária/métodos , Nutrientes/análise , Água/química
2.
Front Immunol ; 14: 1245443, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37771590

RESUMO

Autophagy plays an important role in recognizing and protecting cells from invading intracellular pathogens such as Salmonella. In this work, we investigated the role of p38MAPK/MK2 in modulating the host cell susceptibility to Salmonella infection. Inhibition of p38MAPK or MK2 led to a significant increase of bacterial counts in Salmonella infected mouse embryonic fibroblasts (MEFs), as well as in MK2-deficient (Mk2-/-) cells. Furthermore, western blot analysis showed that Mk2-/- cells have lower level of LC3 lipidation, which is the indicator of general autophagy compared to Mk2-rescued cells. In Mk2-/- cells, we also observed lower activated TANK-binding kinase-1 phosphorylation on Ser172 and p62/SQTM1-Ser403 phosphorylation, which are important to promote the translocation of p62 to ubiquitinated microbes and required for efficient autophagy of bacteria. Furthermore, immunofluorescence analysis revealed reduced colocalization of Salmonella with LC3 and p62 in MEFs. Inhibition of autophagy with bafilomycin A1 showed increased bacterial counts in treated cells compared to control cell. Overall, these results indicate that p38MAPK/MK2-mediated protein phosphorylation modulates the host cell susceptibility to Salmonella infection by affecting the autophagy pathways.


Assuntos
Infecções por Salmonella , Proteínas Quinases p38 Ativadas por Mitógeno , Animais , Camundongos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Fibroblastos/metabolismo , Autofagia
3.
Microorganisms ; 11(8)2023 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-37630684

RESUMO

The yellow early marsh orchid (Dactylorhiza incarnata ssp. ochroleuca) is a critically endangered terrestrial orchid in Britain. Previous attempts to translocate symbiotic seedlings to a site near the last remaining wild site demonstrated some success, with a 10% survival rate despite adverse weather conditions over a two-year period. However, to facilitate future reintroduction efforts or conservation translocations, a more comprehensive understanding of the fungal microbiome and abiotic soil characteristics at the final remaining wild site is required. Obtaining comprehensive information on both the fungal community and soil nutrient composition from wild sites has significant benefits and may prove critical for the success of future conservation translocations involving threatened orchids. This preliminary study, conducted at the last remaining wild site, revealed a significant correlation between the relative abundance of the orchid mycorrhizal fungal order Cantharellales and the concentrations of nitrate and phosphate in the soil. Another orchid mycorrhizal fungal group, Sebacinales, was found to be distributed extensively throughout the site. The composition of fungal communities across the entire site, orchid-hosting and non-orchid-hosting soils is discussed in relation to reinforcing the current population and preventing the extinction of this orchid.

4.
Clin Lymphoma Myeloma Leuk ; 23(11): e379-e385, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37612207

RESUMO

BACKGROUND: Multiple myeloma (MM) is one of the most diagnosed hematologic malignancies in the United States. Despite improvements in therapy, health disparities persist among patients with MM. Here, we aim to determine whether there are disparities in time to diagnosis (TTD) among MM patients with regard to income, race/ethnicity, and gender. PATIENTS: Patients with a monoclonal protein detected in the serum and/or urine and a subsequent bone marrow biopsy confirmed diagnosis of MM were included in the study. METHODS: We extracted data on patients with MM and assessed whether the predictor variables were associated with the primary outcome of TTD, which we define as the time between detection of a monoclonal protein in the serum or urine and bone marrow biopsy diagnosis of MM. RESULTS: Compared to patients with commercial insurance, patients receiving Medicaid (HR: 0.408, 95% CI: 0.206-0.808; P = .010) and patients without insurance (HR: 0.428, 95% CI: 0.207-0.885; P = .022) were significantly more likely to have delayed TTD. TTD was also prolonged if the provider who ordered the testing for the detection of a monoclonal protein was not a hematologist (HR: 0.435, 95% CI: 0.284-0.668; P < .0001). No disparities were found with regard to race/ethnicity or gender. CONCLUSION: This study suggests there may be socioeconomic disparities in TTD among patients with MM. Interventions such as patient navigation may be useful to reduce TTD among socioeconomically disadvantaged patient populations. Further studies need to be conducted to elucidate reasons for delays.


Assuntos
Mieloma Múltiplo , Humanos , Estados Unidos/epidemiologia , Mieloma Múltiplo/diagnóstico , Mieloma Múltiplo/terapia , Disparidades em Assistência à Saúde , Medicaid , Etnicidade , Renda
5.
Brief Funct Genomics ; 2023 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-37461194

RESUMO

We identified 11 conserved stretches in over 6.3 million SARS-CoV-2 genomes including all the major variants of concerns. Each conserved stretch is ≥100 nucleotides in length with ≥99.9% conservation at each nucleotide position. Interestingly, six of the eight conserved stretches in ORF1ab overlapped significantly with well-folded experimentally verified RNA secondary structures. Furthermore, two of the conserved stretches were mapped to regions within the S2-subunit that undergo dynamic structural rearrangements during viral fusion. In addition, the conserved stretches were significantly depleted for zinc-finger antiviral protein (ZAP) binding sites, which facilitated the recognition and degradation of viral RNA. These highly conserved stretches in the SARS-CoV-2 genome were poorly conserved at the nucleotide level among closely related ß-coronaviruses, thus representing ideal targets for highly specific and discriminatory diagnostic assays. Our findings highlight the role of structural constraints at both RNA and protein levels that contribute to the sequence conservation of specific genomic regions in SARS-CoV-2.

6.
Cell Death Discov ; 9(1): 262, 2023 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-37495567

RESUMO

Receptor-interacting protein kinases (RIPK)-1 and -3 play crucial roles in cell fate decisions and are regulated by multiple checkpoint controls. Previous studies have identified IKK1/2- and p38/MK2-dependent checkpoints that phosphorylate RIPK1 at different residues to inhibit its activation. In this study, we investigated TNF-induced death in MAPK-activated protein kinase 2 (MK2)-deficient cells and found that MK2 deficiency or inactivation predominantly leads to necroptotic cell death, even without caspase inhibition. While RIPK1 inhibitors can rescue MK2-deficient cells from necroptosis, inhibiting RIPK3 seems to switch the process to apoptosis. To understand the underlying mechanism of this switch, we screened a library of 149 kinase inhibitors and identified the adenosine analog 5-Iodotubercidin (5-ITu) as the most potent compound that sensitizes MK2-deficient MEFs to TNF-induced cell death. 5-ITu also enhances LPS-induced necroptosis when combined with MK2 inhibition in RAW264.7 macrophages. Further mechanistic studies revealed that 5-ITu induces RIPK1-dependent necroptosis by suppressing IKK signaling in the absence of MK2 activity. These findings highlight the role for the multitarget kinase inhibitor 5-ITu in TNF-, LPS- and chemotherapeutics-induced necroptosis and its potential implications in RIPK1-targeted therapies.

8.
Front Pharmacol ; 14: 1149809, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37007026

RESUMO

Macroautophagy (hereafter referred to as autophagy), a highly conserved metabolic process, regulates cellular homeostasis by degrading dysfunctional cytosolic constituents and invading pathogens via the lysosomal system. In addition, autophagy selectively recycles specific organelles such as damaged mitochondria (via mitophagy), and lipid droplets (LDs; via lipophagy) or eliminates specialized intracellular pathogenic microorganisms such as hepatitis B virus (HBV) and coronaviruses (via virophagy). Selective autophagy, particularly mitophagy, plays a key role in the preservation of healthy liver physiology, and its dysfunction is connected to the pathogenesis of a wide variety of liver diseases. For example, lipophagy has emerged as a defensive mechanism against chronic liver diseases. There is a prominent role for mitophagy and lipophagy in hepatic pathologies including non-alcoholic fatty liver disease (NAFLD), hepatocellular carcinoma (HCC), and drug-induced liver injury. Moreover, these selective autophagy pathways including virophagy are being investigated in the context of viral hepatitis and, more recently, the coronavirus disease 2019 (COVID-19)-associated hepatic pathologies. The interplay between diverse types of selective autophagy and its impact on liver diseases is briefly addressed. Thus, modulating selective autophagy (e.g., mitophagy) would seem to be effective in improving liver diseases. Considering the prominence of selective autophagy in liver physiology, this review summarizes the current understanding of the molecular mechanisms and functions of selective autophagy (mainly mitophagy and lipophagy) in liver physiology and pathophysiology. This may help in finding therapeutic interventions targeting hepatic diseases via manipulation of selective autophagy.

9.
Cytoskeleton (Hoboken) ; 80(7-8): 169-181, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36797225

RESUMO

Septins are cytoskeletal GTPases that form nonpolar filaments and higher-ordered structures and they take part in a wide range of cellular processes. Septins are conserved from yeast to mammals but absent from higher plants. The number of septin genes vary between organisms and they usually form complex heteropolymeric networks. Most septins are known to be capable of GTP hydrolysis which may regulate septin dynamics. Knowledge on regulation of septin function by post-translational modifications is still in its infancy. In this review article, we highlight the post-translational modifications reported for the 13 human septins and discuss their implications on septin functions. In addition to the functionally investigated modifications, we also try to make sense of the complex septin post-translational modification code revealed from large-scale phospho-proteomic datasets. Future studies may determine how these isoform-specific and homology group specific modifications affect septin structure and function.


Assuntos
Proteômica , Septinas , Animais , Humanos , Septinas/metabolismo , Processamento de Proteína Pós-Traducional , Saccharomyces cerevisiae/metabolismo , Mamíferos/metabolismo
10.
Int J Biol Macromol ; 229: 624-635, 2023 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-36587643

RESUMO

Dengue virus (DENV) exploits various cellular pathways including autophagy to assure enhanced virus propagation. The mechanisms of DENV mediated control of autophagy pathway are largely unknown. Our investigations have revealed a novel role for high-mobility group box1 protein (HMGB1) in regulation of cellular autophagy process in DENV-2 infected A549 cell line. While induction of autophagy by rapamycin treatment resulted in enhanced DENV-2 propagation, the blockade of autophagy flux with bafilomycin A1 suppressed viral replication. Furthermore, siRNA-mediated silencing of HMGB1 significantly abrogated dengue induced autophagy, while LPS induced HMGB1 expression counteracted these effects. Interestingly, silencing of HMGB1 showed reduction of BECN1 and stabilization of BCL-2 protein. On the contrary, LPS induction of HMGB1 resulted in enhanced BECN1 and reduction in BCL-2 levels. This study shows that the modulation of autophagy by DENV-2 is HMGB1/BECN1 dependent. In addition, glycyrrhizic acid (GA), a potent HMGB1 inhibitor suppressed autophagy as well as DENV-2 replication. Altogether, our data suggests that HMGB1 induces BECN1 dependent autophagy to promote DENV-2 replication.


Assuntos
Vírus da Dengue , Dengue , Proteína HMGB1 , Humanos , Proteína HMGB1/genética , Lipopolissacarídeos/farmacologia , Replicação Viral , Autofagia , Proteínas Proto-Oncogênicas c-bcl-2 , Dengue/genética
11.
Cell Death Discov ; 9(1): 14, 2023 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-36658119

RESUMO

The TNF receptor-interacting protein kinases (RIPK)-1 and 3 are regulators of extrinsic cell death response pathways, where RIPK1 makes the cell survival or death decisions by associating with distinct complexes mediating survival signaling, caspase activation or RIPK3-dependent necroptotic cell death in a context-dependent manner. Using a mass spectrometry-based screen to find new components of the ripoptosome/necrosome, we discovered the protein-arginine methyltransferase (PRMT)-5 as a direct interaction partner of RIPK1. Interestingly, RIPK3 but not RIPK1 was then found to be a target of PRMT5-mediated symmetric arginine dimethylation. A conserved arginine residue in RIPK3 (R486 in human, R415 in mouse) was identified as the evolutionarily conserved target for PRMT5-mediated symmetric dimethylation and the mutations R486A and R486K in human RIPK3 almost completely abrogated its methylation. Rescue experiments using these non-methylatable mutants of RIPK3 demonstrated PRMT5-mediated RIPK3 methylation to act as an efficient mechanism of RIPK3-mediated feedback control on RIPK1 activity and function. Therefore, this study reveals PRMT5-mediated RIPK3 methylation as a novel modulator of RIPK1-dependent signaling.

12.
Microbiol Spectr ; 10(5): e0273121, 2022 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-35993712

RESUMO

Clinical and in vivo studies have demonstrated a role for hepatitis B virus (HBV)-encoded HBsAg (hepatitis B surface antigen) in HBV-related hepatocellular carcinoma (HCC); however, the underlying mechanisms remain largely unknown. Here, we investigated the role of HBsAg in regulating long noncoding RNAs (lncRNAs) involved in HCC progression. Our analysis of microarray data sets identified LINC00665 as an HBsAg-regulated lncRNA. Furthermore, LINC00665 is upregulated in liver samples from HBV-infected patients as well as in HCC, specifically in HBV-related HCC liver samples. These findings were supported by our in vitro data demonstrating that HBsAg, as well as HBV, positively regulates LINC00665 in multiple HBV cell culture models. Next, we evaluated the oncogenic potential of LINC00665 by its overexpression and CRISPR interference (CRISPRi)-based knockdown in various cell-based assays. LINC00665 promoted cell proliferation, migration, and colony formation but inhibited cell apoptosis in vitro. We then identified the underlying mechanism of HBsAg-mediated regulation of LINC00665. We used immunofluorescence assays to show that HBsAg enhanced the nuclear translocation of NF-κB factors in HepG2 cells, confirming that HBsAg activates NF-κB. Inhibition of NF-κB signaling nullified HBsAg-mediated LINC00665 upregulation, suggesting that HBsAg acts through NF-κB to regulate LINC00665. Furthermore, the LINC00665 promoter contains NF-κB binding sites, and their disruption abrogated HBsAg-induced LINC00665 upregulation. Finally, HBsAg facilitated the enrichment of the NF-κB factors NF-κB1, RelA, and c-Rel in the LINC00665 promoter. Taken together, this work shows that HBsAg can drive hepatocarcinogenesis by upregulating oncogenic LINC000665 through the NF-κB pathway, thereby identifying a novel mechanism in HBV-related HCC. IMPORTANCE Hepatitis B virus (HBV) is a major risk factor for hepatocellular carcinoma (HCC). Numerous reports indicate an oncogenic role for HBV-encoded HBsAg; however, the underlying mechanisms are not well understood. Here, we studied the role of HBsAg in regulating lncRNAs involved in hepatocarcinogenesis. We demonstrate that HBsAg, as well as HBV, positively regulates oncogenic lncRNA LINC00665. The clinical significance of this lncRNA is highlighted by our observation that LINC00665 is upregulated in liver samples during HBV infection and HBV-related HCC. Furthermore, we show LINC00665 can drive hepatocarcinogenesis by promoting cell proliferation, colony formation, and cell migration and inhibiting apoptosis. Taken together, this work identified LINC00665 as a novel gene through which HBsAg can drive hepatocarcinogenesis. Finally, we show that HBsAg enhances LINC00665 levels in hepatocytes by activating the NF-κB pathway, thereby identifying a novel mechanism by which HBV may contribute to HCC.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , RNA Longo não Codificante , Humanos , Vírus da Hepatite B/genética , Vírus da Hepatite B/metabolismo , Antígenos de Superfície da Hepatite B/genética , Antígenos de Superfície da Hepatite B/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia
13.
Front Cell Dev Biol ; 10: 949824, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35784463
14.
Mol Biol Evol ; 39(3)2022 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-35134218

RESUMO

Depletion of CpG dinucleotides in severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) genomes has been linked to virus evolution, host-switching, virus replication, and innate immune responses. Temporal variations, if any, in the rate of CpG depletion during virus evolution in the host remain poorly understood. Here, we analyzed the CpG content of over 1.4 million full-length SARS-CoV-2 genomes representing over 170 million documented infections during the first 17 months of the pandemic. Our findings suggest that the extent of CpG depletion in SARS-CoV-2 genomes is modest. Interestingly, the rate of CpG depletion is highest during early evolution in humans and it gradually tapers off, almost reaching an equilibrium; this is consistent with adaptations to the human host. Furthermore, within the coding regions, CpG depletion occurs predominantly at codon positions 2-3 and 3-1. Loss of ZAP (Zinc-finger antiviral protein)-binding motifs in SARS-CoV-2 genomes is primarily driven by the loss of the terminal CpG within the motifs. Nonetheless, majority of the CpG depletion in SARS-CoV-2 genomes occurs outside ZAP-binding motifs. SARS-CoV-2 genomes selectively lose CpGs-motifs from a U-rich context; this may help avoid immune recognition by TLR7. SARS-CoV-2 alpha-, beta-, and delta-variants of concern have reduced CpG content compared to sequences from the beginning of the pandemic. In sum, we provide evidence that the rate of CpG depletion in virus genomes is not uniform and it greatly varies over time and during adaptations to the host. This work highlights how temporal variations in selection pressures during virus adaption may impact the rate and the extent of CpG depletion in virus genomes.


Assuntos
COVID-19 , SARS-CoV-2 , COVID-19/genética , Genoma Viral , Humanos , Pandemias , SARS-CoV-2/genética , Replicação Viral
15.
Annu Rev Pathol ; 17: 387-402, 2022 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-35073168

RESUMO

Most cancer cases occur in low- and middle-income countries (LMICs). The sophisticated technical and human infrastructure needed for optimal diagnosis, treatment, and monitoring of cancers is difficult enough in affluent countries; it is especially challenging in LMICs. In Western, educated, industrial, rich, democratic countries, there is a growing emphasis on and success with precision medicine, whereby targeted therapy is directed at cancers based on the specific genetic lesions in the cancer. Can such precision approaches be delivered in LMICs? We offer some examples of novel partnerships and creative solutions that suggest that precision medicine may be possible in LMICs given heavy doses of will, creativity, and persistence and a little luck.


Assuntos
Países em Desenvolvimento , Neoplasias , Humanos , Neoplasias/diagnóstico , Neoplasias/genética , Neoplasias/terapia , Pobreza , Medicina de Precisão
16.
Br J Nutr ; 128(5): 888-899, 2022 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34776015

RESUMO

Rice is consumed by nearly half of the global population and a significant source of energy and nutrients. However, rice consumption can also be a significant pathway of inorganic arsenic (iAs) exposure, thus requiring a risk-benefit assessment. This study assessed nutrient element (NE) densities in fifty-five rice types (white, brown and wild rice) marketed in the UK. Densities of essential NE were used to rank rice types in meeting daily NE targets under different consumption scenarios through a newly developed optimisation approach. Using iAs data from these rice types, we assessed the margin of exposure (MOE) for low (the UK) and high (Bangladesh) rice intake scenarios. Our results showed that brown and wild rice are significantly higher in many NE and significantly contribute to dietary reference value (DRV). Our modelling showed that switching to brown or wild rice could increase the intake of several essential nutrients by up to eight times that of white rice. Using rice consumption data for mid-to-high-consumption countries, we estimate that brown rice could provide 100 % adult DRV for Fe, Mg, Cr, P and Mo, and substantial contributions for Zn, Se and K. Our results show that the amount of rice primarily determines risk from iAs consumed rather than the type of rice. Therefore, switching from white to brown or wild rice could be beneficial, provided iAs concentration in rice is within the recommended limits.


Assuntos
Arsênio , Arsenicais , Oryza , Arsênio/análise , Contaminação de Alimentos/análise , Medição de Risco , Nutrientes
17.
Pathogens ; 12(1)2022 Dec 27.
Artigo em Inglês | MEDLINE | ID: mdl-36678391

RESUMO

Hepatitis C virus (HCV) is a bloodborne pathogen that can cause chronic liver disease and hepatocellular carcinoma. The loss of CpGs from virus genomes allows escape from restriction by the host zinc-finger antiviral protein (ZAP). The evolution of HCV in the human host has not been explored in the context of CpG depletion. We analysed 2616 full-length HCV genomes from 1977 to 2021. During the four decades of evolution in humans, we found that HCV genomes have become significantly depleted in (a) CpG numbers, (b) CpG O/E ratios (i.e., relative abundance of CpGs), and (c) the number of ZAP-binding motifs. Interestingly, our data suggests that the loss of CpGs in HCV genomes over time is primarily driven by the loss of ZAP-binding motifs; thus suggesting a yet unknown role for ZAP-mediated selection pressures in HCV evolution. The HCV core gene is significantly enriched for the number of CpGs and ZAP-binding motifs. In contrast to the rest of the HCV genome, the loss of CpGs from the core gene does not appear to be driven by ZAP-mediated selection. This work highlights CpG depletion in HCV genomes during their evolution in humans and the role of ZAP-mediated selection in HCV evolution.

18.
EMBO Rep ; 22(12): e52931, 2021 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-34661963

RESUMO

Aging of hematopoietic stem cells (HSCs) is caused by the elevated activity of the small RhoGTPase Cdc42 and an apolar distribution of proteins. Mechanisms by which Cdc42 activity controls polarity of HSCs are not known. Binder of RhoGTPases proteins (Borgs) are known effector proteins of Cdc42 that are able to regulate the cytoskeletal Septin network. Here, we show that Cdc42 interacts with Borg4, which in turn interacts with Septin7 to regulate the polar distribution of Cdc42, Borg4, and Septin7 within HSCs. Genetic deletion of either Borg4 or Septin7 results in a reduced frequency of HSCs polar for Cdc42 or Borg4 or Septin7, a reduced engraftment potential and decreased lymphoid-primed multipotent progenitor (LMPP) frequency in the bone marrow. Taken together, our data identify a Cdc42-Borg4-Septin7 axis essential for the maintenance of polarity within HSCs and for HSC function and provide a rationale for further investigating the role of Borgs and Septins in the regulation of compartmentalization within stem cells.


Assuntos
Proteínas do Citoesqueleto , Células-Tronco Hematopoéticas , Septinas , Proteínas rho de Ligação ao GTP , Células-Tronco Hematopoéticas/metabolismo , Septinas/genética , Septinas/metabolismo , Transdução de Sinais
19.
Methods Mol Biol ; 2348: 93-111, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34160801

RESUMO

Long noncoding RNAs are defined as transcripts longer than 200 nt with no protein coding potential. Most lncRNAs are expressed in a tissue-specific manner and barring a few, their absolute expression is lower compared to most coding transcripts. Differential expression studies have contributed the most to the functional characterisation of the lncRNAs we know. Sensitive and specific quantification of lncRNA expression is crucial for such studies. SYBR Green dye based real time quantitative PCR is a simple and affordable method of quantitative PCR, wherein the specific binding of the dye to double stranded DNA amplicon emits fluorescence proportionate to the amount of PCR products. Here we describe a detailed protocol for successful lncRNA quantitation by reverse transcription followed by SYBR Green chemistry-based real-time PCR.


Assuntos
Expressão Gênica , RNA Longo não Codificante/genética , Reação em Cadeia da Polimerase em Tempo Real , Linhagem Celular , DNA Primase , Análise de Dados , Humanos , RNA Longo não Codificante/isolamento & purificação , RNA Mensageiro/genética , RNA Mensageiro/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/métodos
20.
Microorganisms ; 9(4)2021 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-33920122

RESUMO

About 90% of Cynorkis species are endemic to the biodiversity hotspot of Madagascar. This terrestrial habitat-specific genus received little study for fungal diversity to support conservation. We evaluated the diversity of culturable fungi of 11 species and soil characteristics from six sites spanning a >40 km radius in and along the region's inselbergs. Peloton-forming fungi were grown in vitro from root/protocorm slices and positively identified using DNA sequencing. The fungal diversity was then correlated with soil pH, NO3-N, P, and K. All species harbored either putative mycorrhizal associates in the Rhizoctonia complex or Hypocreales fungi. Tulasnella Operational Taxonomic Units (OTUs) were most prevalent in all soil types while Serendipita OTUs were found in species inhabiting granite/rock outcrops in moist soil (seepage areas). Most Cynorkis species were present in soil with low NO3-N and P levels with diversity of mycorrhizal fungi inversely correlated to NO3-N levels. Of the different orchid life stages sampled, only one species (Cynorkis fastigiata) yielded putative mycorrhizal fungi from juvenile stages. As diversity of mycorrhizal fungi of Cynorkis spp. was negatively correlated with NO3-N, and majority of the studied taxa were found in soils with low NO3-N and P contents, reintroduction studies must include analysis of N and P in detail. For the first time, we showed that the assemblage of culturable fungi in the roots of habitat-specific species of Cynorkis (Orchidaceae) are intimately tied to specific soil characteristics.

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