The Vaccinium genus in the family Ericaceae comprises many species, including the fruit-bearing blueberry, bilberry, cranberry, huckleberry, and lingonberry. Commercially, the most important are the blueberries (Vaccinium section Cyanococcus), such as Vaccinium corymbosum (northern highbush blueberry), Vaccinium virgatum (rabbiteye blueberry), and Vaccinium angustifolium (lowbush blueberry). The rising popularity of blueberries can partly be attributed to their “superfood” status, with an increasing body of evidence around human health benefits resulting from the fruit metabolites, particularly products of the phenylpropanoid pathway such as anthocyanins. Activation of anthocyanin production by R2R3-MYB transcription factors (TFs) has been characterized in many species, but despite recent studies on blueberry, cranberry, and bilberry, no MYB anthocyanin regulators have been reported for Vaccinium. Indeed, there has been conjecture that at least in bilberry, MYB TFs divergent to the usual type are involved. We report identification of MYBA from blueberry, and show through sequence analysis and functional studies that it is homologous to known anthocyanin-promoting R2R3-MYBs of subgroup 6 of the MYB superfamily. In transient assays, MYBA complemented an anthocyanin MYB mutant of Antirrhinum majus and, together with a heterologous bHLH anthocyanin regulator, activated anthocyanin production in Nicotiana benthamiana. Furthermore anthocyanin accumulation and anthocyanin structural gene expression (assayed by qPCR and RNA-seq analyses) correlated with MYBA expression, and MYBA was able to transactivate the DFR promoter from blueberry and other species. The RNA-seq data also revealed a range of other candidate genes involved in the regulation of anthocyanin production in blueberry fruit. The identification of MYBA will help to resolve the regulatory mechanism for anthocyanin pigmentation in the Vaccinium genus. The sequence information should also prove useful in developing tools for the accelerated breeding of new Vaccinium cultivars.

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Kiwifruit contains the cysteine proteinase actinidin whose strong activity allows kiwifruit to be used as a meat tenderiser. This raises the possibility digestive enzymes, also proteins, are themselves susceptible to degradation by actinidin. Salivary amylase and gastric lipase are exposed to the highest concentrations of actinidin whereas duodenal enzymes are less likely to be inactivated by actinidin due to dilution and inactivation of actinidin by gastric juice. The saliva of six volunteers was exposed to Actinidia deliciosa homogenate and then examined for loss of the starch digesting enzyme, alpha-amylase. In agreement with the known distribution of salivary amylase concentration in saliva, the range of amylase activity within the group of volunteers varied by around 100 fold. Within 5 minutes of incubation of 3 parts saliva to one part green kiwifruit at 37 °C, approximately 85% of the amylase activity was lost. The use of E-64, a selective inhibitor of cysteine proteinases, confirmed that the loss of amylase function was due to actinidin. Amylase protein degradation was followed by SDS-PAGE and western blotting. Recombinant human gastric lipase resisted digestion with kiwifruit even after 30 minutes incubation and remained functionally active after this time period. However, both mountain papaya and pineapple extracts degraded gastric lipase fully during a 30 minutes digestion period. Under conditions where cooked starch is consumed along with kiwifruit it is possible that starch digestion may be retarded whereas lipid digestion in the stomach is unlikely to be affected by kiwifruit consumption.

Highbush blueberry (Vaccinium corymbosum, 2n = 4x = 48) is the most cultivated type of blueberry, both in New Zealand and overseas. Its perceived nutritional value is conferred by phytonutrients, particularly anthocyanins. Identifying the genetic mechanisms that control the biosynthesis of these metabolites would enable faster development of cultivars with improved fruit qualities. Here, we used recently released tools for genetic mapping in autotetraploids to build a high-density linkage map in highbush blueberry and to detect quantitative trait loci (QTLs) for fruit anthocyanin content. Genotyping was performed by target sequencing, with ∼18,000 single nucleotide polymorphism (SNP) markers being mapped into 12 phased linkage groups (LGs). Fruits were harvested when ripe for two seasons and analyzed with high-performance liquid chromatography-mass spectrometry (HPLC-MS): 25 different anthocyanin compounds were identified and quantified. Two major QTLs that were stable across years were discovered, one on LG2 and one on LG4, and the underlying candidate genes were identified. Interestingly, the presence of anthocyanins containing acylated sugars appeared to be under strong genetic control. Information gained in this study will enable the design of molecular markers for marker-assisted selection and will help build a better understanding of the genetic control of anthocyanin biosynthesis in this crop.

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