A tissue-specific protein purification approach in Caenorhabditis elegans identifies novel interaction partners of DLG-1/Discs large

Selma Waaijers, Javier Mu�oz, Christian Berends, Jo�o J. Ramalho, Soenita S. Goerdayal, Low Teck Yew, Adja D. Zoumaro-Djayoon, Michael Hoffmann, Thijs Koorman, Roderick P. Tas, Martin Harterink, Stefanie Seelk, Jana Kerver, Casper C. Hoogenraad, Olaf Bossinger, Baris Tursun, Sander van den Heuvel, Albert J.R. Heck, Mike Boxem

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Background: Affinity purification followed by mass spectrometry (AP/MS) is a widely used approach to identify protein interactions and complexes. In multicellular organisms, the accurate identification of protein complexes by AP/MS is complicated by the potential heterogeneity of complexes in different tissues. Here, we present an in vivo biotinylation-based approach for the tissue-specific purification of protein complexes from Caenorhabditis elegans. Tissue-specific biotinylation is achieved by the expression in select tissues of the bacterial biotin ligase BirA, which biotinylates proteins tagged with the Avi peptide. Results: We generated N- and C-terminal tags combining GFP with the Avi peptide sequence, as well as four BirA driver lines expressing BirA ubiquitously and specifically in the seam and hyp7 epidermal cells, intestine, or neurons. We validated the ability of our approach to identify bona fide protein interactions by identifying the known LGL-1 interaction partners PAR-6 and PKC-3. Purification of the Discs large protein DLG-1 identified several candidate interaction partners, including the AAA-type ATPase ATAD-3 and the uncharacterized protein MAPH-1.1. We have identified the domains that mediate the DLG-1/ATAD-3 interaction, and show that this interaction contributes to C. elegans development. MAPH-1.1 co-purified specifically with DLG-1 purified from neurons, and shared limited homology with the microtubule-associated protein MAP1A, a known neuronal interaction partner of mammalian DLG4/PSD95. A CRISPR/Cas9-engineered GFP::MAPH-1.1 fusion was broadly expressed and co-localized with microtubules. Conclusions: The method we present here is able to purify protein complexes from specific tissues. We uncovered a series of DLG-1 interactors, and conclude that ATAD-3 is a biologically relevant interaction partner of DLG-1. Finally, we conclude that MAPH-1.1 is a microtubule-associated protein of the MAP1 family and a candidate neuron-specific interaction partner of DLG-1.

Original languageEnglish
Article number66
JournalBMC Biology
Volume14
Issue number1
DOIs
Publication statusPublished - 1 Jan 2016
Externally publishedYes

Fingerprint

Caenorhabditis elegans
Purification
purification
Tissue
protein
Proteins
proteins
Biotinylation
Microtubule-Associated Proteins
Neurons
biotinylation
microtubules
Mass Spectrometry
neurons
Clustered Regularly Interspaced Short Palindromic Repeats
Mass spectrometry
Caenorhabditis elegans Proteins
peptide
Peptides
mass spectrometry

Keywords

  • Affinity purification
  • C. elegans
  • Discs large
  • Mass spectrometry
  • Microtubuleassociated
  • Protein complex
  • Tissue-specific

ASJC Scopus subject areas

  • Biotechnology
  • Structural Biology
  • Ecology, Evolution, Behavior and Systematics
  • Physiology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • Plant Science
  • Developmental Biology
  • Cell Biology

Cite this

A tissue-specific protein purification approach in Caenorhabditis elegans identifies novel interaction partners of DLG-1/Discs large. / Waaijers, Selma; Mu�oz, Javier; Berends, Christian; Ramalho, Jo�o J.; Goerdayal, Soenita S.; Teck Yew, Low; Zoumaro-Djayoon, Adja D.; Hoffmann, Michael; Koorman, Thijs; Tas, Roderick P.; Harterink, Martin; Seelk, Stefanie; Kerver, Jana; Hoogenraad, Casper C.; Bossinger, Olaf; Tursun, Baris; van den Heuvel, Sander; Heck, Albert J.R.; Boxem, Mike.

In: BMC Biology, Vol. 14, No. 1, 66, 01.01.2016.

Research output: Contribution to journalArticle

Waaijers, S, Mu�oz, J, Berends, C, Ramalho, JJ, Goerdayal, SS, Teck Yew, L, Zoumaro-Djayoon, AD, Hoffmann, M, Koorman, T, Tas, RP, Harterink, M, Seelk, S, Kerver, J, Hoogenraad, CC, Bossinger, O, Tursun, B, van den Heuvel, S, Heck, AJR & Boxem, M 2016, 'A tissue-specific protein purification approach in Caenorhabditis elegans identifies novel interaction partners of DLG-1/Discs large', BMC Biology, vol. 14, no. 1, 66. https://doi.org/10.1186/s12915-016-0286-x
Waaijers, Selma ; Mu�oz, Javier ; Berends, Christian ; Ramalho, Jo�o J. ; Goerdayal, Soenita S. ; Teck Yew, Low ; Zoumaro-Djayoon, Adja D. ; Hoffmann, Michael ; Koorman, Thijs ; Tas, Roderick P. ; Harterink, Martin ; Seelk, Stefanie ; Kerver, Jana ; Hoogenraad, Casper C. ; Bossinger, Olaf ; Tursun, Baris ; van den Heuvel, Sander ; Heck, Albert J.R. ; Boxem, Mike. / A tissue-specific protein purification approach in Caenorhabditis elegans identifies novel interaction partners of DLG-1/Discs large. In: BMC Biology. 2016 ; Vol. 14, No. 1.
@article{0ff582d299a44bd1a135f5f398244790,
title = "A tissue-specific protein purification approach in Caenorhabditis elegans identifies novel interaction partners of DLG-1/Discs large",
abstract = "Background: Affinity purification followed by mass spectrometry (AP/MS) is a widely used approach to identify protein interactions and complexes. In multicellular organisms, the accurate identification of protein complexes by AP/MS is complicated by the potential heterogeneity of complexes in different tissues. Here, we present an in vivo biotinylation-based approach for the tissue-specific purification of protein complexes from Caenorhabditis elegans. Tissue-specific biotinylation is achieved by the expression in select tissues of the bacterial biotin ligase BirA, which biotinylates proteins tagged with the Avi peptide. Results: We generated N- and C-terminal tags combining GFP with the Avi peptide sequence, as well as four BirA driver lines expressing BirA ubiquitously and specifically in the seam and hyp7 epidermal cells, intestine, or neurons. We validated the ability of our approach to identify bona fide protein interactions by identifying the known LGL-1 interaction partners PAR-6 and PKC-3. Purification of the Discs large protein DLG-1 identified several candidate interaction partners, including the AAA-type ATPase ATAD-3 and the uncharacterized protein MAPH-1.1. We have identified the domains that mediate the DLG-1/ATAD-3 interaction, and show that this interaction contributes to C. elegans development. MAPH-1.1 co-purified specifically with DLG-1 purified from neurons, and shared limited homology with the microtubule-associated protein MAP1A, a known neuronal interaction partner of mammalian DLG4/PSD95. A CRISPR/Cas9-engineered GFP::MAPH-1.1 fusion was broadly expressed and co-localized with microtubules. Conclusions: The method we present here is able to purify protein complexes from specific tissues. We uncovered a series of DLG-1 interactors, and conclude that ATAD-3 is a biologically relevant interaction partner of DLG-1. Finally, we conclude that MAPH-1.1 is a microtubule-associated protein of the MAP1 family and a candidate neuron-specific interaction partner of DLG-1.",
keywords = "Affinity purification, C. elegans, Discs large, Mass spectrometry, Microtubuleassociated, Protein complex, Tissue-specific",
author = "Selma Waaijers and Javier Mu{\"i}¿½oz and Christian Berends and Ramalho, {Jo{\"i}¿½o J.} and Goerdayal, {Soenita S.} and {Teck Yew}, Low and Zoumaro-Djayoon, {Adja D.} and Michael Hoffmann and Thijs Koorman and Tas, {Roderick P.} and Martin Harterink and Stefanie Seelk and Jana Kerver and Hoogenraad, {Casper C.} and Olaf Bossinger and Baris Tursun and {van den Heuvel}, Sander and Heck, {Albert J.R.} and Mike Boxem",
year = "2016",
month = "1",
day = "1",
doi = "10.1186/s12915-016-0286-x",
language = "English",
volume = "14",
journal = "BMC Biology",
issn = "1741-7007",
publisher = "BioMed Central",
number = "1",

}

TY - JOUR

T1 - A tissue-specific protein purification approach in Caenorhabditis elegans identifies novel interaction partners of DLG-1/Discs large

AU - Waaijers, Selma

AU - Mu�oz, Javier

AU - Berends, Christian

AU - Ramalho, Jo�o J.

AU - Goerdayal, Soenita S.

AU - Teck Yew, Low

AU - Zoumaro-Djayoon, Adja D.

AU - Hoffmann, Michael

AU - Koorman, Thijs

AU - Tas, Roderick P.

AU - Harterink, Martin

AU - Seelk, Stefanie

AU - Kerver, Jana

AU - Hoogenraad, Casper C.

AU - Bossinger, Olaf

AU - Tursun, Baris

AU - van den Heuvel, Sander

AU - Heck, Albert J.R.

AU - Boxem, Mike

PY - 2016/1/1

Y1 - 2016/1/1

N2 - Background: Affinity purification followed by mass spectrometry (AP/MS) is a widely used approach to identify protein interactions and complexes. In multicellular organisms, the accurate identification of protein complexes by AP/MS is complicated by the potential heterogeneity of complexes in different tissues. Here, we present an in vivo biotinylation-based approach for the tissue-specific purification of protein complexes from Caenorhabditis elegans. Tissue-specific biotinylation is achieved by the expression in select tissues of the bacterial biotin ligase BirA, which biotinylates proteins tagged with the Avi peptide. Results: We generated N- and C-terminal tags combining GFP with the Avi peptide sequence, as well as four BirA driver lines expressing BirA ubiquitously and specifically in the seam and hyp7 epidermal cells, intestine, or neurons. We validated the ability of our approach to identify bona fide protein interactions by identifying the known LGL-1 interaction partners PAR-6 and PKC-3. Purification of the Discs large protein DLG-1 identified several candidate interaction partners, including the AAA-type ATPase ATAD-3 and the uncharacterized protein MAPH-1.1. We have identified the domains that mediate the DLG-1/ATAD-3 interaction, and show that this interaction contributes to C. elegans development. MAPH-1.1 co-purified specifically with DLG-1 purified from neurons, and shared limited homology with the microtubule-associated protein MAP1A, a known neuronal interaction partner of mammalian DLG4/PSD95. A CRISPR/Cas9-engineered GFP::MAPH-1.1 fusion was broadly expressed and co-localized with microtubules. Conclusions: The method we present here is able to purify protein complexes from specific tissues. We uncovered a series of DLG-1 interactors, and conclude that ATAD-3 is a biologically relevant interaction partner of DLG-1. Finally, we conclude that MAPH-1.1 is a microtubule-associated protein of the MAP1 family and a candidate neuron-specific interaction partner of DLG-1.

AB - Background: Affinity purification followed by mass spectrometry (AP/MS) is a widely used approach to identify protein interactions and complexes. In multicellular organisms, the accurate identification of protein complexes by AP/MS is complicated by the potential heterogeneity of complexes in different tissues. Here, we present an in vivo biotinylation-based approach for the tissue-specific purification of protein complexes from Caenorhabditis elegans. Tissue-specific biotinylation is achieved by the expression in select tissues of the bacterial biotin ligase BirA, which biotinylates proteins tagged with the Avi peptide. Results: We generated N- and C-terminal tags combining GFP with the Avi peptide sequence, as well as four BirA driver lines expressing BirA ubiquitously and specifically in the seam and hyp7 epidermal cells, intestine, or neurons. We validated the ability of our approach to identify bona fide protein interactions by identifying the known LGL-1 interaction partners PAR-6 and PKC-3. Purification of the Discs large protein DLG-1 identified several candidate interaction partners, including the AAA-type ATPase ATAD-3 and the uncharacterized protein MAPH-1.1. We have identified the domains that mediate the DLG-1/ATAD-3 interaction, and show that this interaction contributes to C. elegans development. MAPH-1.1 co-purified specifically with DLG-1 purified from neurons, and shared limited homology with the microtubule-associated protein MAP1A, a known neuronal interaction partner of mammalian DLG4/PSD95. A CRISPR/Cas9-engineered GFP::MAPH-1.1 fusion was broadly expressed and co-localized with microtubules. Conclusions: The method we present here is able to purify protein complexes from specific tissues. We uncovered a series of DLG-1 interactors, and conclude that ATAD-3 is a biologically relevant interaction partner of DLG-1. Finally, we conclude that MAPH-1.1 is a microtubule-associated protein of the MAP1 family and a candidate neuron-specific interaction partner of DLG-1.

KW - Affinity purification

KW - C. elegans

KW - Discs large

KW - Mass spectrometry

KW - Microtubuleassociated

KW - Protein complex

KW - Tissue-specific

UR - http://www.scopus.com/inward/record.url?scp=84991764306&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84991764306&partnerID=8YFLogxK

U2 - 10.1186/s12915-016-0286-x

DO - 10.1186/s12915-016-0286-x

M3 - Article

C2 - 27506200

AN - SCOPUS:84991764306

VL - 14

JO - BMC Biology

JF - BMC Biology

SN - 1741-7007

IS - 1

M1 - 66

ER -