Phylogenetics of the genus Erica and anthocyanin synthesis gene expression in Erica plukenetii

Le Maitre, Nicholas (2017-03)

Thesis (PhD)--Stellenbosch University, 2017.

Thesis

ENGLISH ABSTRACT: The drivers of species radiations are central to questions about the evolution of diversity. The flora of the Cape Floristic Region (CFR) is particularly diverse, has exceptionally high level of endemism and may have radiated at an exceptionally high rate. Various drivers of this radiation have been proposed, including climate change, fire, niche adaptation, persistence of lineages and shifts in pollination syndrome. Erica is the largest genus in the CFR but its radiation has not been well studied phylogenetically. A multiple marker phylogeny would be significant in establishing its radiation rate and further elucidating the role and importance that factors such as biogeography and pollinator shifts, have played in driving its radiation specifically and in the CFR flora in general. Floral colour shifts between red and white flowers have been shown to be important in switches between pollination syndromes. The anthocyanin pathway produces coloured anthocyanins that colour the flowers of plants, also in Erica. A multiple chloroplast and ITS marker region phylogeny was constructed for 597 accessions. Automated and manual alignment strategies were used to generate phylogenies and found to not be significantly different. Overall the phylogeny showed African species are descended from European species and that Mascarean and Drakensberg species may share a common ancestor with Cape species. A single Cape clade is present, sister to one anomalous species, and the sub clades reveal structure primarily related to biogeography and not morphology. Both flower colour and pollination syndrome are highly labile and multiple switches have occurred between anemophily, entomophily and ornithophily. Red flowers and ornithophily have evolved independently on at least 14 occasions. In red flowered Erica plukenetii whole genome sequencing approaches using Illumina NGS sequencing were used to obtain sequences of the anthocyanin pathway genes and their trans-acting regulatory genes. RT-PCR and RT-qPCR were used to measure the expression of these genes in two populations of red-, pink- and white-flowered E. plukenetii. Expression of the CHS and the ANS genes were found to be reduced in white flowers in these populations respectively. Sequencing of the promoter regions of these genes in red-, pink- and white-flowered plants revealed mutations in the promoter binding sites of the white flowered plants that likely are the cause of anthocyanin synthesis enzyme gene down regulation and consequent loss of flower colour. Biogeographical factors and shifts between pollination syndromes that potentially result from changes in red anthocyanin synthesis contributed to the loss of anthocyanin production are therefore likely important drivers of the radiation of Erica in the CFR.

AFRIKAANSE OPSOMMING: Die aandrywers van spesiesverspreiding is sentraal tot vrae oor die evolusie van diversiteit. Die flora van die Kaapse Floristiese Streek (KFS) is veral divers, het 'n baie hoë vlak van endemisme en mag versprei het teen 'n buitengewone hoë tempo. Verskeie aandrywers van hierdie verspreiding word voorgestel, insluitende klimaatsverandering, vuur-, nisaanpassing, volharding van afstammelinge en verskuiwings in bestuiwingsindroom. Erica is die grootste genus in die KFS, maar die verspreiding is nog nie goed bestudeer op filogenetiese vlak nie. ‘n Veelvuldige geen filogenie sou insiggewend wees i.v.m. die bepaling van verspreidingstempos, asook die rol wat biogeografie en bestuiwerverskuiwings op hierdie tempo gehad het in hierdie genus, maar ook in die KFS in geheel. Blommekleurverskuiwings tussen rooi en wit blomme is getoon om belangrik te wees vir die omskakelings tussen bestuiwingsindrome. Die antosianien padweg produseer gekleurde antosianiene wat die blomme van plante kleur, insluitende die van Erica. ‘n Veelvuldige chloroplast en ITS merker filogenie is gegenereer vir 597 versamelinge. Outomatiese asook oplynings met die oog oplyningsstrategieë van DNS volgordes is gebruik om filogenieë te genereer en te vergelyk. Hulle is gevind om nie beduidend te verskil nie. Die filogenie toon dat die Afrikaanse spesies afstam van die Europese spesies en dat Maskareense en Drakensberg spesies 'n gemeenskaplike voorouer met Kaapse spesies deel. 'n Enkele Kaap klade is teenwoordig, die suster van een onreëlmatige spesie, en die sub clades openbaar struktuur wat hoofsaaklik verband hou met biogeografie en nie morfologie nie. Beide blomkleur en bestuiwingsindroom is hoogs labiel en verskeie verskuiwings het plaasgevind tussen wind-, insek- en voëlbestuiwing. Rooi blomme en voëlbestuiwing het onafhanklik ontwikkel op ten minste 14 geleenthede. ‘n Heelgenoom volgorderbepaalingsbenadering was gevolg vir die rooiblom E. plukenetii, op die Illumina platform, in ‘n pogig om die volgordes te bepaal van die antosianien pad gene en hul transwerkende regulerende gene te verkry. RT-PCR en RT-qPCR is gebruik om die uitdrukking van hierdie gene te meet in twee bevolkings van rooi-, pink- en wit-blommende E. plukenetii. Uitdrukking van die CHS en die ANS gene is onderskeidelik gevind om afreguleer te word in wit blomme in hierdie bevolkings. Die promotorstreke se DNS volgordes is vir hierdie monsters bepaal en daar is gevind dat die promotor bindingstreke in die witblom plante gemuteer is. Dit is waarskynlik die oorsaak van antosianiensintese ensiemgene afregulering en gevolglike verlies van blomkleur. Biogeografiese faktore en verskuiwings tussen bestuiwingsindrome wat potensieel as gevolg van veranderinge in rooi antosianien sintese bygedra het tot die verlies van antosianien produksie is dus waarskynlik belangrike drywers van die verspruiding van Erica in die KFS.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/101060
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