Synthetic control devices for gene regulation in Penicillium chrysogenum.
Gene regulation
Hybrid transcription factor
Penicillium chrysogenum
Secondary metabolite production
Synthetic expression system
Synthetic gene cluster
Journal
Microbial cell factories
ISSN: 1475-2859
Titre abrégé: Microb Cell Fact
Pays: England
ID NLM: 101139812
Informations de publication
Date de publication:
18 Nov 2019
18 Nov 2019
Historique:
received:
16
08
2019
accepted:
10
11
2019
entrez:
20
11
2019
pubmed:
20
11
2019
medline:
9
4
2020
Statut:
epublish
Résumé
Orthogonal, synthetic control devices were developed for Penicillium chrysogenum, a model filamentous fungus and industrially relevant cell factory. In the synthetic transcription factor, the QF DNA-binding domain of the transcription factor of the quinic acid gene cluster of Neurospora crassa is fused to the VP16 activation domain. This synthetic transcription factor controls the expression of genes under a synthetic promoter containing quinic acid upstream activating sequence (QUAS) elements, where it binds. A gene cluster may demand an expression tuned individually for each gene, which is a great advantage provided by this system. The control devices were characterized with respect to three of their main components: expression of the synthetic transcription factors, upstream activating sequences, and the affinity of the DNA binding domain of the transcription factor to the upstream activating domain. This resulted in synthetic expression devices, with an expression ranging from hardly detectable to a level similar to that of highest expressed native genes. The versatility of the control device was demonstrated by fluorescent reporters and its application was confirmed by synthetically controlling the production of penicillin. The characterization of the control devices in microbioreactors, proved to give excellent indications for how the devices function in production strains and conditions. We anticipate that these well-characterized and robustly performing control devices can be widely applied for the production of secondary metabolites and other compounds in filamentous fungi.
Sections du résumé
BACKGROUND
BACKGROUND
Orthogonal, synthetic control devices were developed for Penicillium chrysogenum, a model filamentous fungus and industrially relevant cell factory. In the synthetic transcription factor, the QF DNA-binding domain of the transcription factor of the quinic acid gene cluster of Neurospora crassa is fused to the VP16 activation domain. This synthetic transcription factor controls the expression of genes under a synthetic promoter containing quinic acid upstream activating sequence (QUAS) elements, where it binds. A gene cluster may demand an expression tuned individually for each gene, which is a great advantage provided by this system.
RESULTS
RESULTS
The control devices were characterized with respect to three of their main components: expression of the synthetic transcription factors, upstream activating sequences, and the affinity of the DNA binding domain of the transcription factor to the upstream activating domain. This resulted in synthetic expression devices, with an expression ranging from hardly detectable to a level similar to that of highest expressed native genes. The versatility of the control device was demonstrated by fluorescent reporters and its application was confirmed by synthetically controlling the production of penicillin.
CONCLUSIONS
CONCLUSIONS
The characterization of the control devices in microbioreactors, proved to give excellent indications for how the devices function in production strains and conditions. We anticipate that these well-characterized and robustly performing control devices can be widely applied for the production of secondary metabolites and other compounds in filamentous fungi.
Identifiants
pubmed: 31739777
doi: 10.1186/s12934-019-1253-3
pii: 10.1186/s12934-019-1253-3
pmc: PMC6859608
doi:
Substances chimiques
Fungal Proteins
0
Transcription Factors
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
203Subventions
Organisme : FP7 People: Marie-Curie Actions
ID : 607332
Organisme : H2020 Marie Skłodowska-Curie Actions
ID : 713482
Références
Nucleic Acids Res. 2016 Sep 30;44(17):e136
pubmed: 27325743
Cell. 1984 Dec;39(3 Pt 2):499-509
pubmed: 6096007
Microb Cell Fact. 2009 Jun 04;8:31
pubmed: 19497126
Fungal Genet Biol. 2006 Nov;43(11):727-38
pubmed: 16843015
Appl Environ Microbiol. 2010 Nov;76(21):7109-15
pubmed: 20851974
Nat Biotechnol. 2008 Oct;26(10):1161-8
pubmed: 18820685
Folia Microbiol (Praha). 2010 Mar;55(2):126-32
pubmed: 20490754
Nat Methods. 2012 Mar 11;9(4):391-5
pubmed: 22406855
J Mol Biol. 1985 Sep 5;185(1):65-81
pubmed: 3900423
Appl Microbiol Biotechnol. 1999 Aug;52(2):196-207
pubmed: 10499259
Appl Environ Microbiol. 2017 Dec 15;84(1):null
pubmed: 29079620
Genetics. 2010 Oct;186(2):735-55
pubmed: 20697123
Fungal Genet Biol. 2016 Apr;89:72-83
pubmed: 26555930
Nat Methods. 2015 Mar;12(3):219-22, 5 p following 222
pubmed: 25581800
Nucleic Acids Res. 2018 Oct 12;46(18):e111
pubmed: 29924368
PLoS One. 2016 Feb 22;11(2):e0148320
pubmed: 26901642
ACS Synth Biol. 2016 Jul 15;5(7):754-64
pubmed: 27072635
ACS Chem Biol. 2018 Nov 16;13(11):3193-3205
pubmed: 30339758
Microb Cell Fact. 2016 Nov 11;15(1):192
pubmed: 27835989
Dev Biol. 2010 Mar 15;339(2):225-9
pubmed: 19682982
Methods Mol Biol. 2012;835:1-16
pubmed: 22183644
Biotechnol Bioeng. 2012 Nov;109(11):2884-95
pubmed: 22565375
Nucleic Acids Res. 2014 Apr;42(6):e48
pubmed: 24445804
Front Microbiol. 2015 Mar 16;6:184
pubmed: 25852654
ACS Synth Biol. 2014 Mar 21;3(3):129-39
pubmed: 24299494
Nature. 1988 Oct 6;335(6190):563-4
pubmed: 3047590
ACS Synth Biol. 2015 Sep 18;4(9):975-86
pubmed: 25871405
Crit Rev Microbiol. 2010 May;36(2):146-67
pubmed: 20210692
ACS Synth Biol. 2017 Mar 17;6(3):471-484
pubmed: 27973777
Appl Environ Microbiol. 2011 May;77(9):2975-83
pubmed: 21378046
Mol Cell Biol. 1991 Nov;11(11):5746-55
pubmed: 1922075
Cell. 2010 Apr 30;141(3):536-48
pubmed: 20434990
Microb Cell Fact. 2014 Apr 11;13:53
pubmed: 24725602
Methods. 2014 Apr 1;66(3):433-40
pubmed: 23792917
Nat Commun. 2016 Oct 03;7:13010
pubmed: 27694947
Appl Environ Microbiol. 2012 Oct;78(19):7107-13
pubmed: 22865068
Nat Prod Rep. 2013 Aug;30(8):1121-38
pubmed: 23832108
Appl Environ Microbiol. 2005 Feb;71(2):672-8
pubmed: 15691916
Proc Natl Acad Sci U S A. 2015 Mar 3;112(9):2847-52
pubmed: 25691737
Funct Integr Genomics. 2009 May;9(2):167-84
pubmed: 19156454
BMC Microbiol. 2005 Jan 13;5:1
pubmed: 15649330
BMC Genomics. 2015 Nov 14;16:937
pubmed: 26572918
Biochem J. 2009 Jan 15;417(2):467-76
pubmed: 18834333
Anal Biochem. 2003 Jul 1;318(1):152-4
pubmed: 12782044
Nature. 2009 Mar 19;458(7236):362-6
pubmed: 19092803
Nat Protoc. 2013 Nov;8(11):2281-2308
pubmed: 24157548
BMC Genomics. 2009 Feb 10;10:75
pubmed: 19203396
Crit Rev Biotechnol. 2000;20(1):17-48
pubmed: 10770226
Methods Mol Biol. 2018;1772:213-232
pubmed: 29754231
Microb Cell Fact. 2019 Apr 2;18(1):63
pubmed: 30940138
Fungal Genet Biol. 2016 Apr;89:62-71
pubmed: 26701309
Methods Mol Biol. 2016;1478:53-78
pubmed: 27730575
Fungal Biol Biotechnol. 2016 Aug 31;3:6
pubmed: 28955465
Microbiol Rev. 1985 Sep;49(3):338-58
pubmed: 2931582
ACS Synth Biol. 2015 May 15;4(5):625-33
pubmed: 25226362