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Evidence of 4-Cl-IAA and IAA Bound to Proteins in Pea Fruit and Seeds

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Abstract

The auxins 4-chloroindole-3-acetic acid (4-Cl-IAA) and indole-3-acetic acid (IAA) occur naturally in pea vegetative and fruit tissues (Pisum sativum L.). Previous work has shown that 4-Cl-IAA can substitute for the seeds in the stimulation of pea pericarp growth, whereas IAA is ineffective. Both auxins are found as free acids and as low-molecular-weight conjugates from organic solvent-soluble extracts from pea fruit. Here we present evidence for an additional conjugated auxin species that was not soluble in organic solvent and yielded 4-Cl-IAA and IAA after strong alkaline hydrolysis, suggestive of auxin attachment to pea seed and pericarp proteins. The solvent-insoluble conjugated 4-Cl-IAA in young pericarp was on average 15-fold greater than solvent-soluble 4-Cl-IAA. The solvent-insoluble conjugated IAA was approximately half the levels reported for the solvent-soluble IAA fraction. To identify putative 4-Cl-IAA-bound proteins, polyclonal antibodies were raised to 4-Cl-IAA linked to bovine serum albumin protein (BSA). Immunoblots probed with anti-4-Cl-IAA-BSA antiserum detected three to four unique bands (32–40 kDa) in primarily maternal tissues, and a different set of protein bands were detected in mainly embryonic tissues (ca. 65–74 kDa in mature seed). 4-Cl-IAA and IAA were also identified from protein fractions separated by polyacrylamide gel electrophoresis using GC-MS. These data show that the majority of 4-Cl-IAA, the growth-active auxin in young pea pericarp, and significant levels of IAA are linked to protein fractions. Auxin-proteins may function in regulation of free bioactive 4-Cl-IAA and IAA levels, and/or 4-Cl-IAA or IAA may be targeted to specific proteins post-translationally to modify protein function or stability.

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Acknowledgments

We thank Thomas P. Krick (Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota) for the MALDI-TOF spectrum. The authors acknowledge funding support for this research from the Natural Sciences and Engineering Research Council of Canada to JAO, and financial support to JDC from the U.S. National Science Foundation (IBN 0111530 and IOS 0820940), the University of Minnesota Grant-in-Aid program, and the Gordon and Margaret Bailey Endowment for Environmental Horticulture.

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Authors and Affiliations

  1. Department of Horticultural Science and Microbial and Plant Genomics Institute, University of Minnesota, St. Paul, MN, 55108, USA

    Seijin Park & Jerry D. Cohen

  2. Plant Physiology and Molecular Biology Research Group, Department of Agricultural, Food and Nutritional Science, University of Alberta, 4-10 Agriculture/Forestry Centre, Edmonton, AB, T6G 2P5, Canada

    Jocelyn A. Ozga & Dennis M. Reinecke

  3. Joint Center for Biosciences, 7-45, Songdo-dong, Yeonsu-ku, Incheon, 406-840, Korea

    Seijin Park

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  1. Seijin Park
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  2. Jocelyn A. Ozga
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Correspondence to Jocelyn A. Ozga.

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Park, S., Ozga, J.A., Cohen, J.D. et al. Evidence of 4-Cl-IAA and IAA Bound to Proteins in Pea Fruit and Seeds. J Plant Growth Regul 29, 184–193 (2010). https://doi.org/10.1007/s00344-009-9123-6

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  • DOI: https://doi.org/10.1007/s00344-009-9123-6

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