Abstract
The economic value of a black walnut (Juglans nigra L.) tree is strongly determined by the quality and quantity of darkly colored heartwood in its stem. To understand the regulation of heartwood formation, we analyzed the region of heartwood formation in walnut stems (i.e., the transition zone, TZ) for the expression of 80 ESTs. Semi-quantitative RT-PCR and real-time PCR was performed to detect expres- sion changes of candidate genes in the TZ and sap- wood of trees harvested in summer and fall. Results revealed that the transcript of a clone containing two presumed EF-hand motifs was expressed at higher levels in the TZ than in other xylem tissues. Analysis of the full-length coding sequence revealed that the black walnut gene JnCML-like is similar to grancal- cin-like calcium-binding EF hand proteins in Arabi- dopsis thaliana (At3g10300) and Zea mays (NM 001153810). A model of the predicted structure of JnCML-like showed it is similar to grancalcin and m-calpain, penta-EF-hand family proteins associated with cell proliferation, differentiation and program- med cell death. JnCML-like transcript was detected in tissue from the region of the pith meristem, and in roots, embryogenic callus, vascular cambium, female flowers, male flowers, green leaves, and partially and fully senescent leaves of black walnut, although tran- script abundance varied considerably among these tissues.
Highlights
Calcium is a critical second messenger in all eukaryotic cells responding to external stimuli
A model of the predicted structure of JnCML-like showed it is similar to grancalcin and m-calpain, penta-EF-hand family proteins associated with cell proliferation, differentiation and programmed cell death
In order to investigate the expression of JnCML-like in the TZ versus sapwood in more detail, the sapwood portion of black walnut logs harvested in summer and fall were divided into three zones: an interior sapwood zone, which was next to the transition zone; a middle sapwood zone; and an exterior sapwood zone, close to the cambium (Figure 1)
Summary
Calcium is a critical second messenger in all eukaryotic cells responding to external stimuli. Calcium interacts with a large number of proteins, but among the most studied are proteins containing the EF-hand motif. Calcium-dependent EF-hand motif-containing proteins are common in animals, plants, and fungi [1,2]. The basic EF-hand motif is highly conserved and usually consists of two perpendicular 10 to 12 residue alpha helices with a 12-residue loop region. Together these three domains form a single helix-loop-helix calcium-binding site. Two hundred and fifty genes encoding EF-hand-containing proteins have been identified in Arabidopsis thaliana [4]. Group I proteins are found in plants and fungi and have a clear role in regulating development, including apoptosis [6]. The best characterized phytocalpain, Defective Kernel 1 (DEK1), is described as calpainlike [7]; it appears to have a complex role in the regulation of plant growth and development
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