TY - JOUR
T1 - A novel panel of yeast assays for the assessment of thiamin and its biosynthetic intermediates in plant tissues
AU - Strobbe, Simon
AU - Verstraete, Jana
AU - Fitzpatrick, Teresa B.
AU - Faustino, Maria
AU - Lourenço, Tiago F.
AU - Oliveira, M. Margarida
AU - Stove, Christophe
AU - Van Der Straeten, Dominique
N1 - Funding Information:
The authors would like to express their gratitude to Prof. David Shintani and Prof. Ming-Hsiun Hsieh for providing them with the tpk1/tpk2 double mutant and pale green1 mutant, respectively. Furthermore, the authors are very thankful to Dr Raymond Wightman for providing the RWY16 strain. DVDS and CS are grateful to Ghent University for financial support (GOA 01G00409; Bijzonder Onderzoeksfonds UGent). SS was indebted to the Agency for Innovation by Science and Technology in Flanders (IWT) for a predoctoral fellowship. SS gratefully acknowledges the Bijzonder Onderzoeksfonds (BOF-UGent) for a postdoctorial fellowship (BOF.P-DO.2019.0008.01). JV would like to thank the Research Foundation Flanders (FWO) for granting her a PhD fellowship (1S61617N). TBF expresses gratitude to the Swiss National Science Foundation for funding (grants 31003A-141117/1 and 31003A_162555/1). MMO would like to acknowledge ‘FCT – Fundação para a Ciência e a Tecnologia, I.P.’ through R&D Unit GREEN-IT Bioresources for Sustainability (UIDB/04551/2020 and UIDP/04551/2020) TFL acknowledges contract CEECIND/03641/2017. MF is grateful for PhD fellowship PD/BD/148694/2019. Authors declare no conflict of interest.
Funding Information:
The authors would like to express their gratitude to Prof. David Shintani and Prof. Ming‐Hsiun Hsieh for providing them with the double mutant and mutant, respectively. Furthermore, the authors are very thankful to Dr Raymond Wightman for providing the RWY16 strain. DVDS and CS are grateful to Ghent University for financial support (GOA 01G00409; Bijzonder Onderzoeksfonds UGent). SS was indebted to the Agency for Innovation by Science and Technology in Flanders (IWT) for a predoctoral fellowship. SS gratefully acknowledges the Bijzonder Onderzoeksfonds (BOF‐UGent) for a postdoctorial fellowship (BOF.P‐DO.2019.0008.01). JV would like to thank the Research Foundation Flanders (FWO) for granting her a PhD fellowship (1S61617N). TBF expresses gratitude to the Swiss National Science Foundation for funding (grants 31003A‐141117/1 and 31003A_162555/1). MMO would like to acknowledge ‘FCT – Fundação para a Ciência e a Tecnologia, I.P.’ through R&D Unit GREEN‐IT Bioresources for Sustainability (UIDB/04551/2020 and UIDP/04551/2020) TFL acknowledges contract CEECIND/03641/2017. MF is grateful for PhD fellowship PD/BD/148694/2019. Authors declare no conflict of interest. tpk1/tpk2 pale green1
Publisher Copyright:
© 2022 The Authors New Phytologist © 2022 New Phytologist Foundation.
PY - 2022/4
Y1 - 2022/4
N2 - Thiamin (or thiamine), known as vitamin B1, represents an indispensable component of human diets, being pivotal in energy metabolism. Thiamin research depends on adequate vitamin quantification in plant tissues. A recently developed quantitative liquid chromatography–tandem mass spectrometry (LC–MS/MS) method is able to assess the level of thiamin, its phosphorylated entities and its biosynthetic intermediates in the model plant Arabidopsis thaliana, as well as in rice. However, their implementation requires expensive equipment and substantial technical expertise. Microbiological assays can be useful in deter-mining metabolite levels in plant material and provide an affordable alternative to MS-based analysis. Here, we evaluate, by comparison to the LC–MS/MS reference method, the potential of a carefully chosen panel of yeast assays to estimate levels of total vitamin B1, as well as its biosynthetic intermediates pyrimidine and thiazole in Arabidopsis samples. The examined panel of Saccharomyces cerevisiae mutants was, when implemented in microbiological assays, capable of correctly assigning a series of wild-type and thiamin biofortified Arabidopsis plant samples. The assays provide a readily applicable method allowing rapid screening of vitamin B1 (and its biosynthetic intermediates) content in plant material, which is particularly useful in metabolic engineering approaches and in germplasm screening across or within species.
AB - Thiamin (or thiamine), known as vitamin B1, represents an indispensable component of human diets, being pivotal in energy metabolism. Thiamin research depends on adequate vitamin quantification in plant tissues. A recently developed quantitative liquid chromatography–tandem mass spectrometry (LC–MS/MS) method is able to assess the level of thiamin, its phosphorylated entities and its biosynthetic intermediates in the model plant Arabidopsis thaliana, as well as in rice. However, their implementation requires expensive equipment and substantial technical expertise. Microbiological assays can be useful in deter-mining metabolite levels in plant material and provide an affordable alternative to MS-based analysis. Here, we evaluate, by comparison to the LC–MS/MS reference method, the potential of a carefully chosen panel of yeast assays to estimate levels of total vitamin B1, as well as its biosynthetic intermediates pyrimidine and thiazole in Arabidopsis samples. The examined panel of Saccharomyces cerevisiae mutants was, when implemented in microbiological assays, capable of correctly assigning a series of wild-type and thiamin biofortified Arabidopsis plant samples. The assays provide a readily applicable method allowing rapid screening of vitamin B1 (and its biosynthetic intermediates) content in plant material, which is particularly useful in metabolic engineering approaches and in germplasm screening across or within species.
KW - biofortification
KW - metabolic engineering
KW - microbiological assays
KW - nutritional improvement
KW - Saccharomyces cerevisiae
KW - turbidimetry
KW - vitamin quantification
UR - http://www.scopus.com/inward/record.url?scp=85124544408&partnerID=8YFLogxK
U2 - 10.1111/nph.17974
DO - 10.1111/nph.17974
M3 - Article
C2 - 35037254
AN - SCOPUS:85124544408
SN - 0028-646X
VL - 234
SP - 748
EP - 763
JO - New Phytologist
JF - New Phytologist
IS - 2
ER -