Metabolism under the spotlight in senescence

Abstract

Mammals and poikilotherms share twomelanized pigment cell types, the mela-nocytes derived from the neural crestand the retinal pigmented epithelium(RPE) derived from the brain (Bhartiet al., 2006). Zebrafish, chicken andhuman studies have identified some ofthe main genes involved in melanin pro-duction in the RPE; however, the extentto which the transcriptome of thesetwo melanized cell types is sharedremains unclear. In addition to the mel-anized cells, fish and other poikilo-therms have other pigment cellsderived from the neural crest, includingiridophores. The characteristic shinysilver appearance of the iridophores isdue to their cytoplasmic reflecting plate-lets, membrane-bound organelles con-taining guanine crystals that areorganized at tightly regulated fixed dis-tances from each other to generate theiridescence by a thin-layer interferencemechanism (Bagnara and Matsumoto,2006). We still do not know how theseplatelets are organized, and how thesecells synthesize abundant guanine forthe platelets while maintaining enoughpurines for nucleic acid production. Thecharacterization of the transcriptomesof iridophores and melanocytes (bothtypes) by Higdon et al. sheds light onthe genes regulating the interestingproperties of the iridophore, as well asthe similarities between the melano-cytes.Higdon et al. developed a method toisolate populations of each of threepigment cells – RPE, and neural crest-derived melanocytes and iridophores– from zebrafish embryos. Cells wereisolated from 3 days post-fertilizationembryos by density gradient centrifuga-tion, using the differences in density ofmelanin and guanine to separate pig-ment cells from the cells of the rest ofthe embryo. To isolate the RPE, embry-onic eyes were dissected, disaggregat-ed and then separated by densitygradient centrifugation. NC-derived mel-anocytes and iridophores were isolatedby density gradient centrifugation ofwhole disaggregated embryos afterremoval of eyes, followed by takingadvantage of the reflective properties ofiridophores to use FACS to separateiridophores from melanocytes. Indepen-dent pooled samples of each pigmentcell were isolated, cDNA extracted andthe transcriptomes sequenced by RNA-seq. Unfortunately, for technical rea-sons, their method was unsuitable forisolating the fourth zebrafish pigmentcell type, the xanthophore.Higdon et al. first aimed to identifygenes shared between the three pig-ment cells, which might indicate sharedfunctions of all pigment cell types. Theyidentified 28 genes, including someencoding ribosomal proteins, and geneswhose role in any pigment cell remainsunclear and will require further study(crfb5 or ghitm). One surprise in this listwas the zebrafish albino gene (slc45a2),which is interesting because albino fishhave reported no iridophore defect; theauthors suggest that it may have a rolein pH regulation in reflecting plateletsas well as in melanosomes.A comparison of genes sharedbetween pairs of pigment cell typesreveals shared functions, but also islikely to reflect shared developmentalorigin. Unsurprisingly, they found 214genes enriched in both melanocytesand RPE, but not iridophores. As bothcells produce the same pigment, mela-nocytes and RPE co-express manygenes involved in melanin synthesisand melanosome regulation, such asdct, tyrp1, pah and tyrp1b. They alsoshare transcription factors from theforkhead box and homeobox-containinggroups (foxo1b, foxp4, hmx1, otx1a,etc.). More unexpected, however, wasthe finding that the 38 most highlyexpressed genes in both melanocytesand RPE are also expressed (albeit atmuch lower levels) in iridophores. Theyalso identified 62 genes enriched inmelanocytes and iridophores, butnot RPE. The authors note that thesefindings likely reflect the shared originsof melanocytes and iridophores fromthe neural crest, because the lists ofshared genes include several well-known NC and pigment cell regulatorsand markers (sox10, crestin, mc1r andthe cell adhesion molecule pcdh10a).Moreover, some transcription factorsthat have not been studied previously,such as foxo1a or cdk15, were alsoenriched in both pigment cells and willwarrant further study using targetedgene mutagenesis. Only one gene wasenriched in RPE and iridophores, whichwas expected because they do notshare a close developmental origin orpigment type.Finally, Higdon et al. have identifiedgenes specific to each cell type. Theiranalysis identified 108 genes specificallyenriched in melanocytes, and 24 in theRPE. However, the most interestingresults concern the characterization ofthe iridophore transcriptome, which pro-vides insights into the genetics underly-ing these highly specialized cells.The authors identify 346 genes whoseexpression is significantly enriched iniridophores. However, by filtering the listto include only highly enriched genes (atleast 30-fold greater than melanocytesand RPE, and 100-fold greater thanwhole embryos), they highlight 30 geneslikely to play major roles in iridophorespecialization. This list includes genes,such as slc23 l, that may act as a guan-ine transporter, although whether thisacts in the plasma membrane or in thereflecting platelet membrane remains tobe determined. More surprisingly, thelist also includes gpnmb, encoding atransmembrane glycoprotein previouslydescribed in melanocytes as both aplasma membrane protein and a mela-nosome component that regulates theexpression of melanosomal proteins andmelanosome formation; the authorsspeculate that it may have a similar func-tion in the generation of the iridophorereflecting platelet organelle.One striking finding of their study isthe enrichment of transcripts encodingenzymes involved in the purine synthe-sis pathway, from extracellular glucoseimport and glycolysis, through de novosynthesis and purine retrieval. Upregula-tion of certain genes in the purineCoverage on: Higdon, C.W., Mitra, R.D.,and Johnson, S.L. (2013). Gene expres-sion analysis of zebrafish melanocytes,iridophores, and retinal pigmented epi-thelium reveals indicators of biologicalfunction and developmental origin.PLoS ONE 8(7), e67801. doi:10.1371/journal.pone.0067801