Biology subjects

Green, B. D.

Publications and source records attributed to Green, B. D..

4 recordsLinked to original sources

bioRxiv2024

The oncogene SLC35F2 is a high-specificity transporter for the micronutrients queuine and queuosine

The nucleobase queuine (q) and its nucleoside queuosine (Q) are micronutrients derived from bacteria that are acquired from the gut microbiome and/or diet in humans. Following cellular uptake, Q is incorporated at the wobble base (position 34) of tRNAs with a GUN anticodon, which is important for efficient translation. Early studies suggested that cytosolic uptake of queuine is mediated by a selective transporter that is regulated by mitogenic signals, but the identity of this transporter has remained elusive. Here, through a cross-species bioinformatic search and genetic validation, we have identified the solute carrier family member SLC35F2 as a unique transporter for both queuine and queuosine in Schizosaccharomyces pombe and Trypanosoma brucei. Furthermore, gene disruption in HeLa cells revealed that SLC35F2 is the sole transporter for queuosine in HeLa cells (Km 174 nM) and a high-affinity transporter for the queuine nucleobase (Km 67 nM), with the presence of another low-affinity transporter (Km 259 nM) in these cells. Competition uptake studies show that SLC35F2 is not a general transporter for other canonical ribonucleobases or ribonucleosides, but selectively imports q and Q. The identification of SLC35F2, an oncogene, as the transporter of both q and Q advances our understanding of how intracellular levels of queuine and queuosine are regulated and how their deficiency contributes to a variety of pathophysiological conditions, including neurological disorders and cancer. Significance StatementThe discovery of SLC35F2 as the eukaryotic transporter of queuine and queuosine is key to understanding how these micronutrients are salvaged from the human gut and distributed to different body tissues. Queuosine modification of tRNAs enhances the accuracy and efficiency of codon-anticodon pairing and regulates a range of biological and pathophysiological states, including oxidative stress responses, cancer, learning, memory, and gut homeostasis.

biochemistry↗

bioRxiv2024

Development, validation and application of an LC-MS/MS method quantifying free forms of the micronutrients queuine and queuosine in human plasma using a surrogate matrix approach

Queuosine (Q) is a hypermodified 7-deaza-guanosine nucleoside exclusively synthesized by bacteria. This micronutrient and its respective nucleobase form queuine (q) are salvaged by humans either from gut microflora, or digested food. Depletion of Q-tRNA in human or mouse cells causes protein misfolding that triggers endoplasmic reticular stress and activation of the unfolded protein responses. In vivo, this reduces neuronal architecture of the mouse brain affecting learning and memory. Herein, a sensitive method for quantifying free q and Q in human blood was developed, optimised and validated. After evaluating q/Q extraction efficiency in several different solid-phase sorbents, Bond Elut PBA (phenylboronic acid) cartridges were found to have the highest extraction recovery for q (82%) and Q (71%) from pooled human plasma. PBS with 4% BSA was used as surrogate matrix for method development and validation. An LC-MS/MS method was validated across the concentration range of 0.0003 - 1 {micro}M for both q and Q, showing excellent: linearity (r2 = 0.997 (q) and r2 = 0.998 (Q)), limit of quantification (0.0003 {micro}M), accuracy (100.39% - 125.71%) and precision (CV% < 15.68%). In a sampling of healthy volunteers (n = 44) there was no significant difference in q levels between male (n = 14; mean = 0.0068 {micro}M) and female (n = 30; mean = 0.0080 {micro}M) participants (p = 0.50). Q was not detected in human plasma. This validated method can now be used to further our understanding of the role of q/Q in nutrition, physiology and pathology.

biochemistry↗

bioRxiv2022

Eight week dietary inclusion of nitrite-containing frankfurter exacerbates colorectal cancer pathology, increases oxidative stress, alters metabolism and causes gut dybiosis in APCmin mice

Colorectal cancer (CRC) is the second most prevelant malignancy in Europe and diet is an important modifiable risk factor. Processed meat consumption, including meats with preservative salts such as sodium nitrite, have been implicated in CRC pathogenesis. This study investigated how the CRC pathology and metabolic status of adenomatous polyposis coli (APC) multiple intestinal neoplasia (min) mice was perturbed following 8 weeks of pork meat consumption. Dietary inclusions (15%) of either nitrite-free pork, nitrite-free sausage or nitrite-containing sausage (frankfurter) were compared against a parallel control group (100% chow). Comprehensive studies investigated: gastrointestinal tract histology (tumours, aberrant crypt foci (ACF) and mucin deplin foci (MDF), lipid peroxidation (urine and serum), faecal microbiota and serum metabolomics (599 metabolites). After 8 weeks mice consuming the frankfurter diet had 53% more (P=0.014) gastrointestinal tumours than control, although ACF and MDF did not differ. Urine and serum lipid peroxidation markers were 59% (P=0.001) and 108% (P=0.001) higher, respectively in the frankfurter group. Gut dysbiosis was evident in these mice with comparably fewer Bacteriodes and more Firmicutes. Fasting serum levels of trimethylamine N-oxide (TMAO) and numerous triglycerides were elevated. Various serum phosphotidylcholine species were decreased. These results demonstrate that nitrite-containing sausages may exaccerbate the development of CRC pathology in APCMin mice to a greater extent than nitrite-free sausages, and this is associated with greater lipid peroxidation, wide-ranging metabolic alternation and gut dysbiosis.

biochemistry↗

bioRxiv2020

Impact of industrial production system parameters on chicken microbiomes: mechanisms to improve performance and reduce Campylobacter

BackgroundThe factors affecting host-pathogen ecology in terms of the microbiome remain poorly studied. Chickens are a key source of protein with gut health heavily dependent on the complex microbiome which has key roles in nutrient assimilation and vitamin and amino acid biosynthesis. The chicken gut microbiome may be influenced by extrinsic production system parameters such as Placement Birds/m2 (stocking density), feed type and additives. Such parameters, in addition to on-farm biosecurity may influence performance and also pathogenic bacterial numbers such as Campylobacter. In this study, three different production systems Normal (N), Higher Welfare (HW) and Omega-3 Higher Welfare (O) were investigated "in a natural environment" at day 7 and day 30 with a range of extrinsic parameters assessing performance in correlation with microbial dynamics and Campylobacter presence. ResultsOur data identified production system N as significantly dissimilar from production systems HW and O when comparing the prevalence of genera. An increase in Placement Birds/m2 density led to a decrease in environmental pressure influencing the microbial community structure. Prevalence of genera such as Eisenbergiella within HW and O, and likewise Alistipes within N were representative. These genera have roles directly relating to energy metabolism, amino acid, nucleotide and short chain fatty acid (SCFA) utilisation. Thus, an association exists between consistent and differentiating parameters of the production systems, that affect feed utilisation, advance our knowledge of mechanistic underpinnings, leading to competitive exclusion of genera based on competition for nutrients and other factors. Campylobacter was identified within specific production system and presence was linked with the increased diversity and increased environmental pressure on microbial community structure. Addition of Omega-3 though did alter prevalence of specific genera, in our analysis did not differentiate itself from HW production system. However, Omega-3 was linked with a positive impact on weight gain. ConclusionsOverall, our results show that microbial communities in different industrial production systems are deterministic in elucidating the underlying biological confounders, and these recommendations are transferable to farm practices and diet manipulation leading to improved performance and better intervention strategies against Campylobacter within the food chain. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=125 SRC="FIGDIR/small/084251v2_ufig1.gif" ALT="Figure 1"> View larger version (63K): org.highwire.dtl.DTLVardef@c43951org.highwire.dtl.DTLVardef@8efa5corg.highwire.dtl.DTLVardef@d6db70org.highwire.dtl.DTLVardef@1e09fb9_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗