Nodule-specific PLAT domain proteins are expanded in the Medicago lineage and required for nodulation

Diana I. Trujillo, Kevin A.T. Silverstein, Nevin D. Young

Research output: Contribution to journalArticlepeer-review

21 Scopus citations

Abstract

Symbiotic nitrogen fixation in legumes is mediated by an interplay of signaling processes between plant hosts and rhizobial symbionts. In legumes, several secreted protein families have undergone expansions and play key roles in nodulation. Thus, identifying lineage-specific expansions (LSEs) of nodulation-associated genes can be a strategy to discover candidate gene families. Using bioinformatic tools, we identified 13 LSEs of nodulation-related secreted protein families, each unique to either Glycine, Arachis or Medicago lineages. In the Medicago lineage, nodule-specific Polycystin-1, Lipoxygenase, Alpha Toxin (PLAT) domain proteins (NPDs) expanded to five members. We examined NPD function using CRISPR/Cas9 multiplex genome editing to create Medicago truncatula NPD knockout lines, targeting one to five NPD genes. Mutant lines with differing combinations of NPD gene inactivations had progressively smaller nodules, earlier onset of nodule senescence, or ineffective nodules compared to the wild-type control. Double- and triple-knockout lines showed dissimilar nodulation phenotypes but coincided in upregulation of a DHHC-type zinc finger and an aspartyl protease gene, possible candidates for the observed disturbance of proper nodule function. By postulating that gene family expansions can be used to detect candidate genes, we identified a family of nodule-specific PLAT domain proteins and confirmed that they play a role in successful nodule formation.

Original languageEnglish (US)
Pages (from-to)1538-1550
Number of pages13
JournalNew Phytologist
Volume222
Issue number3
DOIs
StatePublished - May 2019

Bibliographical note

Funding Information:
We would like to thank Colby Starker, for kindly sharing the rhizobial reporter strain used in this study. We would further like to thank Liana Burghardt for valuable advice in RNA-seq analysis, Peter Tiffin for assistance with experimental design, and Shaun Curtin and Tomá≤s C≤ermák for their expertise and guidance in Medicago transformation and CRISPR methodology. Finally, we would like to thank Catalina Pislariu for sharing her ongoing findings in separate research on NPD genes and advice regarding NPD experiments. Computational analyses were conducted using resources from the Minnesota Supercomputing Institute. Research was funded by National Science Foundation grant IOS-1237993 and the Minnesota Agricultural Experiment Station grant MN22-082.

Funding Information:
We would like to thank Colby Starker, for kindly sharing the rhizobial reporter strain used in this study. We would further like to thank Liana Burghardt for valuable advice in RNA-seq analysis, Peter Tiffin for assistance with experimental design, and Shaun Curtin and Tom?? ?erm?k for their expertise and guidance in Medicago transformation and CRISPR methodology. Finally, we would like to thank Catalina Pislariu for sharing her ongoing findings in separate research on NPD genes and advice regarding NPD experiments. Computational analyses were conducted using resources from the Minnesota Supercomputing Institute. Research was funded by National Science Foundation grant IOS-1237993 and the Minnesota Agricultural Experiment Station grant MN22-082.

Publisher Copyright:
© 2019 The Authors. New Phytologist © 2019 New Phytologist Trust

Keywords

  • lineage-specific expansion
  • multiplex genome editing
  • nodulation-related secreted proteins
  • nodule organogenesis
  • nodule-specific PLAT domain

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