Characterization of a calcium/calmodulin-dependent protein kinase homolog from maize roots showing light-regulated gravitropism

Abstract

Roots of many species respond to gravity (gravitropism) and grow downward only if illuminated. This light-regulated root gravitropism is phytochrome-dependent, mediated by calcium, and inhibited by KN-93, a specific inhibitor of calcium/calmodulin-dependent protein kinase II (CaMK II). A cDNA encoding MCK1, a maize homolog of mammalian CaMK, has been isolated from roots of maize (Zea mays L.). The MCK1 gene is expressed in root tips, the site of perception for both light and gravity. Using the [35S]CaM gel-overlay assay we showed that calmodulin-binding activity of the MCK1 is abolished by 50 microM KN-93, but binding is not affected by 5 microM KN-93, paralleling physiological findings that light-regulated root gravitropism is inhibited by 50 microM KN-93, but not by 5 microM KN-93. KN-93 inhibits light-regulated gravitropism by interrupting transduction of the light signal, not light perception, suggesting that MCK1 may play a role in transducing light. This is the first report suggesting a physiological function for a CaMK homolog in light signal transduction.

Inactivation of Plasma Membrane–Localized CDPK-RELATED KINASE5 Decelerates PIN2 Exocytosis and Root Gravitropic Response inArabidopsis

“…For example, KN-93, an inhibitor of animal calmodulinactivated kinase II, was found to abolish light-regulated root gravitropism in maize (Zea mays) and to compete effectively with CaM binding of the maize CRK MCK1 (Lu et al, 1996). Nonetheless, specific roles of CRKs in the control of root gravitropism remained thus far hidden because the available inhibitors and agonists could target more than one classes of signaling factors.…”

“…All so far known CRKs, including CRK5, share a conserved C-terminal calmodulin (CaM) binding domain, which overlaps with the kinase autoinhibitory domain (Zhang et al, 2002). Selective CaM binding in the presence of Ca 2+ simulates autophosphorylation but leads only to marginal increase of substrate phosphorylation by CRKs, which are equally active without CaM in the presence of either Ca 2+ or the Ca 2+ chelator EGTA (Lindzen and Choi, 1995;Furumoto et al, 1996;Lu et al, 1996;Wang et al, 2001;Zhang et al, 2002;Hua et al, 2003Hua et al, , 2004Wang et al, 2004;Leclercq et al, 2005).…”

Gravity, light and plant form

“…The redistribution' of auxin is generally considered to be responsible for the differentia] growth that gives rise to gravitropism. However, various second messengers, such as calcium and calmodulin, have also been shown to be involved in gravitropism (Hasenstein & Evans 1986;Roux & Serlin 1987;Bethke et al 1995;Lu et al 1996). The mechanism of polar auxin transport has been the focus of much research and a general model, known as the chemiosmotic hypothesis of polar auxin transport, has received general acceptance.…”

Cellular localization and biochemical characterization of a novel calcium-dependent protein kinase from tobacco

“…W38 using ZAPcDNA Synthesis Kit, following the manufacturer's instructions (Invitrogene). The library was then screened using maize MCK1 cDNA [11] as a probe, and a positive plasmid containing NtCPK5, named pNtCPK5 was isolated and sequenced.…”

“…Phylogenic tree analyses indicate that NtCPK5 subfamily including NtCPK4 and StCPK1 is more related to CRK than to CPK. The sequence identity between NtCPK5 and CRK proteins such as maize MCK1 (accession number 1839597) [11] and from pBIm [17] and GFP from pBI101-GFP [17] into pUC18 in frame to produce C-terminal GFP tagged fusion protein. After sequence confirmation, these vectors were bombarded into the epidermal cells of onion.…”