Ca2+ functions as a second messenger in all eukaryotes. Such Ca2+ signaling is used to coordinate plant responses to numerous stimuli, both developmental and environmental. Ca2+ signals are often transduced via the action of the ubiquitous Ca2+ sensor calmodulin (CaM). CaM-dependent protein phosphorylation forms an important component of such signal transduction pathways, including that regulating the initiation and development of symbiotic rhizobial nodules in legumes such as soybean (Glycine max). To further understand the role of Ca2+/CaM during nodule organogenesis, a nodule cDNA expression library was screened using radiolabeled CaM as a probe to identify novel CaM-binding proteins (CaMBPs). This screen resulted in the identification of a previously uncharacterized receptor-like kinase, termed GmCaMK1. The CaM-binding domain (CaMBD) of GmCaMK1 is located in a 24 residue region of GmCaMK1, which overlaps with the subdomain XI of a conserved Ser/Thr kinase domain. This CaMBD bound CaM in a Ca2+-dependent manner, and with high affinity (Kd = 1.4 nM). Furthermore, two hydrophobic residues (V372 and L375) were identified as critical for GmCaMK1-CaM interaction. Recombinant GmCaMK1 exhibited protein kinase activity in vitro, with autophosphorylation activity unaffected by the presence or absence of Ca2+/CaM. GmCaMK1 expression is enriched in developing nodules and main roots, and highest expression level was observed in lateral roots. While the function of CaM-binding to GmCaMK1 remains unclear, the affinity and Ca2+-dependence of the GmCaMK1-CaM interaction strongly suggests that GmCaMK1 is a physiologically relevant CaM target. The Arabidopsis ortholog of GmCaMK1, AtCaMK1 also bound CaM when expressed as a recombinant protein. GmCaMK1 is part of a multi-member family in soybean, as are putative homologs across taxa, suggesting that this is a novel, conserved family of CaMBPs. / Thesis (Master, Biology) -- Queen's University, 2010-01-28 16:00:48.69
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:OKQ.1974/5424 |
| Date | 03 February 2010 |
| Creators | DEFALCO, THOMAS A |
| Contributors | Queen's University (Kingston, Ont.). Theses (Queen's University (Kingston, Ont.)) |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English, English |
| Detected Language | English |
| Type | Thesis |
| Format | 4499944 bytes, application/pdf |
| Rights | This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner. |
| Relation | Canadian theses |
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