Integrative analysis of physiology and genomics provides insights into freeze tolerance adaptations of Acacia koa along an elevational cline.

ABSTRACT

Natural selection for plant species in heterogeneous environments creates genetic variation for traits such as cold tolerance. While physiological or molecular analyses have been used to evaluate stress tolerance adaptations, combining these approaches may provide deeper insight. Acacia koa (koa) occurs from sea level to 2300 m in Hawai'i, USA. At high elevations, natural koa populations have declined due to deforestation, and freeze tolerance is a limiting factor for tree regeneration. We used physiology and molecular analyses to evaluate cold tolerance of koa populations from low (300-750 m), middle (750-1500 m), and high elevations (1500-2100 m). Half of the seedlings were cold acclimated by exposure to progressively lowered air temperatures for eight weeks (from 25.6/22.2°C to 8/4°C, day/night). Using the whole plant physiology-freezing test and koa C-repeat Binding Factor CBF genes, our results indicated that koa can be cold-acclimated when exposed to low, non-freezing temperatures. Seedlings from high elevations had consistently higher expression of Koa CBF genes associated with cold tolerance, helping to explain variation in cold-hardy phenotypes. Evaluation of the genetic background of 22 koa families across the elevations with low coverage RNA sequencing indicated that high elevation koa had relatively low values of heterozygosity, suggesting that adaptation is more likely to arise in the middle and low elevation sources. This physiology and molecular data for cold tolerance of koa across the elevation gradient of the Hawaiian Islands provides insights into natural selection processes and may help to support guidelines for conservation and seed transfer in forest restoration efforts.