JnCML-like, an EF-hand motif-containing gene seasonally upregulated in the transition zone of black walnut (Juglans nigra L.)

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.