Protoplasma DOI 10.1007/s00709-017-1147-3 ORIGINAL ARTICLE Kingdom Chromista and its eight phyla: a new synthesis emphasising periplastid protein targeting, cytoskeletal and periplastid evolution, and ancient divergences Thomas Cavalier-Smith1 Received: 12 April 2017 /Accepted: 18 July 2017 # The Author(s) 2017. This article is an open access publication Abstract In 1981 I established kingdom Chromista, distin- membranes generates periplastid vesicles that fuse with the guished from Plantae because of its more complex arguably derlin-translocon-containing periplastid reticulum chloroplast-associated membrane topology and rigid tubular (putative red algal trans-Golgi network homologue; present multipartite ciliary hairs. Plantae originated by converting a in all chromophytes except dinoflagellates). I explain chromist cyanobacterium to chloroplasts with Toc/Tic translocons; origin from ancestral corticates and neokaryotes, reappraising most evolved cell walls early, thereby losing phagotrophy. tertiary symbiogenesis; a chromist cytoskeletal synapomor- Chromists originated by enslaving a phagocytosed red alga, phy, a bypassing microtubule band dextral to both centrioles, surrounding plastids by two extra membranes, placing them favoured multiple axopodial origins. I revise chromist higher within the endomembrane system, necessitating novel protein classification by transferring rhizarian subphylum Endomyxa import machineries. Early chromists retained phagotrophy, from Cercozoa to Retaria; establishing retarian subphylum remaining naked and repeatedly reverted to heterotrophy by Ectoreta for Foraminifera plus Radiozoa, apicomonad sub- losing chloroplasts. Therefore, Chromista include secondary classes, new dinozoan classes Myzodinea (grouping phagoheterotrophs (notably ciliates, many dinoflagellates, Colpovora gen. n., Psammosa), Endodinea, Sulcodinea, and Opalozoa, Rhizaria, heliozoans) or walled osmotrophs subclass Karlodinia; and ranking heterokont Gyrista as phy- (Pseudofungi, Labyrinthulea), formerly considered protozoa lum not superphylum. or fungi respectively, plus endoparasites (e.g. Sporozoa) and all chromophyte algae (other dinoflagellates, chromeroids, Keywords Chromist periplastid membrane . Chloroplast ochrophytes, haptophytes, cryptophytes). I discuss their ori- protein targeting . Chromist periplastid reticulum . gin, evolutionary diversification, and reasons for making Microtubularcentriolarroots .Chromistevolution .Sporozoan chromists one kingdom despite highly divergent cytoskele- conoid origin tons and trophic modes, including improved explanations for periplastid/chloroplast protein targeting, derlin evolution, and ciliary/cytoskeletal diversification. I conjecture that transit- Introduction: chromist importance and aims of this peptide-receptor-mediated ‘endocytosis’ from periplastid paper Chromista is one of five eukaryotic kingdoms recognised in a Handling Editor: Ulrich Kutschera comprehensive seven-kingdom classification of life Electronic supplementary material The online version of this article (Ruggiero et al. 2015). As here critically reassessed, (https://doi.org/10.1007/s00709-017-1147-3) contains supplementary Chromista comprise eight distinctive phyla, not just three as material, which is available to authorized users. in the first substantial systematic treatment 30 years ago — * Thomas Cavalier-Smith (Cavalier-Smith 1986) 5 years after Chromista was [email protected] established (Cavalier-Smith 1981a). Chromista have turned out to include the vast majority of marine algae and of hetero- 1 Department of Zoology, University of Oxford, South Parks Road, trophic protists, whether marine or in soil or freshwater, and Oxford OX1 3PS, UK some of the most serious human disease agents such as T. Cavalier-Smith malaria parasites and agricultural pathogens like potato blight different paralogues and chromist nuclei and nucleomorphs and sugar beet rhizomania disease, making chromists im- (relict enslaved red algal nuclei) kept different red algal derlin mensely important for ocean ecology, soil biology, climate paralogues for periplastid protein targeting. My discussions on stability, agriculture, and medicine, as well as for fundamental cytoskeletal and ciliary evolution, though rather detailed in understanding of eukaryote evolution and biodiversity. They places, are set in the broad context of overall eukaryote cyto- have a greater range in radically different body plans and skeletal evolution and therefore include some wider implica- lifestyles than the entire plant kingdom and more phyla than tions for eukaryote cell evolution and cell biology in general. kingdoms Fungi or Protozoa. Only animals and bacteria have For convenient reference in a complex field, I summarise an more phyla than chromists, but even they cannot match improved higher-level classification of chromists; by remov- chromists in their remarkable range of contrasting adaptive ing a few past confusions, its revisions enable new cell evo- zones—from giant brown algal kelps longer than a blue whale lutionary insights. As the paper is long, I highlight 15 major to ciliates like Paramecium, dinoflagellates that power coral novel conclusions at the end. reefs or kill shellfish, the most abundant predators in soil (sarcomonad Cercozoa), parasites like Toxoplasma whose cysts are allegedly lodged in a third of human brains and Distinction of Chromista from Plantae Plasmodium that causes malaria, diatoms whose silica frus- tules were once essential for making dynamite or polishing In 1981 kingdom Plantae of Haeckel (1866)—equivalent to astronomical telescope mirrors, and foraminifera or kingdom Vegetabilia or Regnum Vegetabile of Linnaeus haptophyte plankton like Emiliania that can be seen from (1767)—was restricted to all eukaryotes having plastids locat- outer space and made the white cliffs of Dover with their ed in the cytosol that originated directly from an internally calcareous scales and are probably the most speciose photo- enslaved cyanobacterium from which they inherited an enve- synthetic oceanic flagellates and exude volatile chemicals that lope of only two membranes (Cavalier-Smith 1981a). Plantae affect cloud formation and global energy balance. comprise subkingdoms Viridiplantae (green plants), using There are probably in excess of 150,000 free-living chlorophyll b as an accessory photosynthetic pigment, and chromist species, the most speciose being diatoms (estimated Biliphyta (red algae and glaucophytes) that retained at ~100,000 species) and foraminifera (~10,000 living and ~ phycobilisomes from the ancestral cyanobacterial endosymbi- 40,000 fossil species), many thousands undescribed. Parasitic ont instead (Cavalier-Smith 1982, 1998). The key steps in the chromists could be ten times that, as chromist Sporozoa prob- symbiogenetic origin of chloroplasts from cyanobacteria were ably infect every insect and every other animal species, and evolution of membrane transporters for exporting photosyn- other chromists to infect numerous plants, and even some thetic products and machinery for importing nuclear-coded protozoa or other chromists. Already named chromist species proteins (Cavalier-Smith 1982, 2000a, 2013a). Later, multiple (over 180,000; Corliss 2000) may be only the tip of the ice- gene transfers from the enslaved cyanobacterium into the nu- berg. There are probably far more species of chromist than of cleus and losses of the bacterial cell wall were secondary— plants or protozoa, conceivably even more than fungi, and peptidoglycan being retained in chloroplast envelopes of certainly more individual chromists than plants and animals glaucophytes and basal streptophyte Viridiplantae (lost three combined. Possibly, only viruses and bacteria exceed them in times in plant evolution: in red algae, thus absent also in numbers. What are their distinctive features? Why were they chromists; in Chlorophyta; and in the fern/seed plant clade) established as a kingdom separate from Plantae, Fungi, and (Hirano et al. 2016). As predicted (Cavalier-Smith 1982), Protozoa, where they were once misclassified? chloroplasts of all Plantae share an evolutionarily homologous This paper answers both questions in the next four sections protein import machinery (Toc for import across their outer and then provides a new synthesis aimed to better establish membrane (OM) which evolved from the cyanobacterial OM chromist evolutionary unity, clarify their origin, and outline by replacing its outer leaflet lipopolysaccharide by host phos- how their major lineages evolved from a shared ancestral body phatidylcholine (PC) and Tic for traversing their inner mem- plan. Two major innovations are a radically revised interpre- brane; Bölter and Soll 2016). This shared machinery (modi- tation of chromist chloroplast membrane evolution and pro- fied from cyanobacterial protein export machinery) and the tein targeting, including correcting widespread misconcep- fact that chloroplast DNA multigene trees group all chloro- tions about the character and very limited evolutionary role plasts as a single subclade of cyanobacteria (Ochoa de Alda of tertiary symbiogenesis, and thorough reevaluation of et al. 2014) led to general acceptance that chloroplasts origi- centriolar root evolution and evolutionary diversification of nated only once, and Plantae as redefined in 1981 are ciliary transition zones across the kingdom, relating both to monophyletic. innovations in cell motility and feeding and to phylogenetic Chromophyte algae (those using chlorophyll