In vivo protein trapping produces a functional expression codex of the vertebrate proteome

Karl J. Clark; Darius Balciunas; Hans Martin Pogoda; Yonghe Ding; Stephanie E. Westcot; Victoria M. Bedell; Tammy M. Greenwood; Mark D. Urban; Kimberly J. Skuster; Andrew M. Petzold; Jun Ni; Aubrey L. Nielsen; Ashok Patowary; Vinod Scaria; Sridhar Sivasubbu; Xiaolei Xu; Matthias Hammerschmidt; Stephen C. Ekker

doi:10.1038/nmeth.1606

In vivo protein trapping produces a functional expression codex of the vertebrate proteome

Karl J. Clark, Darius Balciunas, Hans Martin Pogoda, Yonghe Ding, Stephanie E. Westcot, Victoria M. Bedell, Tammy M. Greenwood, Mark D. Urban, Kimberly J. Skuster, Andrew M. Petzold, Jun Ni, Aubrey L. Nielsen, Ashok Patowary, Vinod Scaria, Sridhar Sivasubbu, Xiaolei Xu, Matthias Hammerschmidt, Stephen C. Ekker

Research output: Contribution to journal › Article › peer-review

132 Scopus citations

Abstract

We describe a conditional in vivo protein-trap mutagenesis system that reveals spatiotemporal protein expression dynamics and can be used to assess gene function in the vertebrate Danio rerio. Integration of pGBT-RP2.1 (RP2), a gene-breaking transposon containing a protein trap, efficiently disrupts gene expression with >97% knockdown of normal transcript amounts and simultaneously reports protein expression for each locus. The mutant alleles are revertible in somatic tissues via Cre recombinase or splice-site-blocking morpholinos and are thus to our knowledge the first systematic conditional mutant alleles outside the mouse model. We report a collection of 350 zebrafish lines that include diverse molecular loci. RP2 integrations reveal the complexity of genomic architecture and gene function in a living organism and can provide information on protein subcellular localization. The RP2 mutagenesis system is a step toward a unified 'codex' of protein expression and direct functional annotation of the vertebrate genome.

Original language	English (US)
Pages (from-to)	506-512
Number of pages	7
Journal	Nature Methods
Volume	8
Issue number	6
DOIs	https://doi.org/10.1038/nmeth.1606
State	Published - Jun 2011

Bibliographical note

Funding Information:
The US National Institute on Drug Abuse (DA14546), US National Institute of General Medical Sciences (GM63904), National Institute of Diabetes and Digestive and Kidney Diseases (F30DK083219 and P30DK084567) and the Mayo Foundation provided funding for this research. S.S. and V.S. acknowledge funding support from the Council of Scientific and Industrial Research (grant FAC002), India. We thank members of the Center for Genome Engineering at the University of Minnesota for providing collaborative discussion and resources, the staff of the Zebrafish Core Facility at the Mayo Clinic for providing zebrafish care, D. Argue for programming the zfishbook website, E. Klee for bioinformatic analyses of the zebrafish transcriptome used in theoretical design and testing of the RP2 system, A. Person for assistance in injection of fluorescently labeled dextrans, and InSciEd Out participants and Summer Undergraduate Research Fellow, N. Boczek, for imaging RFP-expressing fish.

Publisher link

10.1038/nmeth.1606

Find It

Find It at U of M Libraries

Cite this

Clark, K. J., Balciunas, D., Pogoda, H. M., Ding, Y., Westcot, S. E., Bedell, V. M., Greenwood, T. M., Urban, M. D., Skuster, K. J., Petzold, A. M., Ni, J., Nielsen, A. L., Patowary, A., Scaria, V., Sivasubbu, S., Xu, X., Hammerschmidt, M., & Ekker, S. C. (2011). In vivo protein trapping produces a functional expression codex of the vertebrate proteome. Nature Methods, 8(6), 506-512. https://doi.org/10.1038/nmeth.1606

Clark, KJ, Balciunas, D, Pogoda, HM, Ding, Y, Westcot, SE, Bedell, VM, Greenwood, TM, Urban, MD, Skuster, KJ, Petzold, AM, Ni, J, Nielsen, AL, Patowary, A, Scaria, V, Sivasubbu, S, Xu, X, Hammerschmidt, M & Ekker, SC 2011, 'In vivo protein trapping produces a functional expression codex of the vertebrate proteome', Nature Methods, vol. 8, no. 6, pp. 506-512. https://doi.org/10.1038/nmeth.1606

@article{5b32f28c16674ce0a5b89c04b0ac7fff,

title = "In vivo protein trapping produces a functional expression codex of the vertebrate proteome",

abstract = "We describe a conditional in vivo protein-trap mutagenesis system that reveals spatiotemporal protein expression dynamics and can be used to assess gene function in the vertebrate Danio rerio. Integration of pGBT-RP2.1 (RP2), a gene-breaking transposon containing a protein trap, efficiently disrupts gene expression with >97% knockdown of normal transcript amounts and simultaneously reports protein expression for each locus. The mutant alleles are revertible in somatic tissues via Cre recombinase or splice-site-blocking morpholinos and are thus to our knowledge the first systematic conditional mutant alleles outside the mouse model. We report a collection of 350 zebrafish lines that include diverse molecular loci. RP2 integrations reveal the complexity of genomic architecture and gene function in a living organism and can provide information on protein subcellular localization. The RP2 mutagenesis system is a step toward a unified 'codex' of protein expression and direct functional annotation of the vertebrate genome.",

author = "Clark, {Karl J.} and Darius Balciunas and Pogoda, {Hans Martin} and Yonghe Ding and Westcot, {Stephanie E.} and Bedell, {Victoria M.} and Greenwood, {Tammy M.} and Urban, {Mark D.} and Skuster, {Kimberly J.} and Petzold, {Andrew M.} and Jun Ni and Nielsen, {Aubrey L.} and Ashok Patowary and Vinod Scaria and Sridhar Sivasubbu and Xiaolei Xu and Matthias Hammerschmidt and Ekker, {Stephen C.}",

note = "Funding Information: The US National Institute on Drug Abuse (DA14546), US National Institute of General Medical Sciences (GM63904), National Institute of Diabetes and Digestive and Kidney Diseases (F30DK083219 and P30DK084567) and the Mayo Foundation provided funding for this research. S.S. and V.S. acknowledge funding support from the Council of Scientific and Industrial Research (grant FAC002), India. We thank members of the Center for Genome Engineering at the University of Minnesota for providing collaborative discussion and resources, the staff of the Zebrafish Core Facility at the Mayo Clinic for providing zebrafish care, D. Argue for programming the zfishbook website, E. Klee for bioinformatic analyses of the zebrafish transcriptome used in theoretical design and testing of the RP2 system, A. Person for assistance in injection of fluorescently labeled dextrans, and InSciEd Out participants and Summer Undergraduate Research Fellow, N. Boczek, for imaging RFP-expressing fish.",

year = "2011",

month = jun,

doi = "10.1038/nmeth.1606",

language = "English (US)",

volume = "8",

pages = "506--512",

journal = "Nature Methods",

issn = "1548-7091",

number = "6",

}

TY - JOUR

T1 - In vivo protein trapping produces a functional expression codex of the vertebrate proteome

AU - Clark, Karl J.

AU - Balciunas, Darius

AU - Pogoda, Hans Martin

AU - Ding, Yonghe

AU - Westcot, Stephanie E.

AU - Bedell, Victoria M.

AU - Greenwood, Tammy M.

AU - Urban, Mark D.

AU - Skuster, Kimberly J.

AU - Petzold, Andrew M.

AU - Ni, Jun

AU - Nielsen, Aubrey L.

AU - Patowary, Ashok

AU - Scaria, Vinod

AU - Sivasubbu, Sridhar

AU - Xu, Xiaolei

AU - Hammerschmidt, Matthias

AU - Ekker, Stephen C.

N1 - Funding Information: The US National Institute on Drug Abuse (DA14546), US National Institute of General Medical Sciences (GM63904), National Institute of Diabetes and Digestive and Kidney Diseases (F30DK083219 and P30DK084567) and the Mayo Foundation provided funding for this research. S.S. and V.S. acknowledge funding support from the Council of Scientific and Industrial Research (grant FAC002), India. We thank members of the Center for Genome Engineering at the University of Minnesota for providing collaborative discussion and resources, the staff of the Zebrafish Core Facility at the Mayo Clinic for providing zebrafish care, D. Argue for programming the zfishbook website, E. Klee for bioinformatic analyses of the zebrafish transcriptome used in theoretical design and testing of the RP2 system, A. Person for assistance in injection of fluorescently labeled dextrans, and InSciEd Out participants and Summer Undergraduate Research Fellow, N. Boczek, for imaging RFP-expressing fish.

PY - 2011/6

Y1 - 2011/6

N2 - We describe a conditional in vivo protein-trap mutagenesis system that reveals spatiotemporal protein expression dynamics and can be used to assess gene function in the vertebrate Danio rerio. Integration of pGBT-RP2.1 (RP2), a gene-breaking transposon containing a protein trap, efficiently disrupts gene expression with >97% knockdown of normal transcript amounts and simultaneously reports protein expression for each locus. The mutant alleles are revertible in somatic tissues via Cre recombinase or splice-site-blocking morpholinos and are thus to our knowledge the first systematic conditional mutant alleles outside the mouse model. We report a collection of 350 zebrafish lines that include diverse molecular loci. RP2 integrations reveal the complexity of genomic architecture and gene function in a living organism and can provide information on protein subcellular localization. The RP2 mutagenesis system is a step toward a unified 'codex' of protein expression and direct functional annotation of the vertebrate genome.

AB - We describe a conditional in vivo protein-trap mutagenesis system that reveals spatiotemporal protein expression dynamics and can be used to assess gene function in the vertebrate Danio rerio. Integration of pGBT-RP2.1 (RP2), a gene-breaking transposon containing a protein trap, efficiently disrupts gene expression with >97% knockdown of normal transcript amounts and simultaneously reports protein expression for each locus. The mutant alleles are revertible in somatic tissues via Cre recombinase or splice-site-blocking morpholinos and are thus to our knowledge the first systematic conditional mutant alleles outside the mouse model. We report a collection of 350 zebrafish lines that include diverse molecular loci. RP2 integrations reveal the complexity of genomic architecture and gene function in a living organism and can provide information on protein subcellular localization. The RP2 mutagenesis system is a step toward a unified 'codex' of protein expression and direct functional annotation of the vertebrate genome.

UR - http://www.scopus.com/inward/record.url?scp=79957812969&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79957812969&partnerID=8YFLogxK

U2 - 10.1038/nmeth.1606

DO - 10.1038/nmeth.1606

M3 - Article

C2 - 21552255

AN - SCOPUS:79957812969

SN - 1548-7091

VL - 8

SP - 506

EP - 512

JO - Nature Methods

JF - Nature Methods

IS - 6

ER -

In vivo protein trapping produces a functional expression codex of the vertebrate proteome

Abstract

Bibliographical note

Publisher link

Find It

Other files and links

Fingerprint

Cite this