Quantitative proteomics of the tobacco pollen tube secretome identifies novel pollen tube guidance proteins important for fertilization

List of primers used in this study. (XLSX 12 kb

Comprehensive analysis of tobacco pollen transcriptome unveils common pathways in polar cell expansion and underlying heterochronic shift during spermatogenesis

<p>Abstract</p> <p>Background</p> <p>Many flowering plants produce bicellular pollen. The two cells of the pollen grain are destined for separate fates in the male gametophyte, which provides a unique opportunity to study genetic interactions that govern guided single-cell polar expansion of the growing pollen tube and the coordinated control of germ cell division and sperm cell fate specification. We applied the Agilent 44 K tobacco gene chip to conduct the first transcriptomic analysis of the tobacco male gametophyte. In addition, we performed a comparative study of the Arabidopsis root-hair trichoblast transcriptome to evaluate genetic factors and common pathways involved in polarized cell-tip expansion.</p> <p>Results</p> <p>Progression of pollen grains from freshly dehisced anthers to pollen tubes 4 h after germination is accompanied with > 5,161 (14.9%) gametophyte-specific expressed probes active in at least one of the developmental stages. In contrast, > 18,821 (54.4%) probes were preferentially expressed in the sporophyte. Our comparative approach identified a subset of 104 pollen tube-expressed genes that overlap with root-hair trichoblasts. Reverse genetic analysis of selected candidates demonstrated that Cu/Zn superoxide dismutase 1 (CSD1), a WD-40 containing protein (BP130384), and Replication factor C1 (NtRFC1) are among the central regulators of pollen-tube tip growth. Extension of our analysis beyond the second haploid mitosis enabled identification of an opposing-dynamic accumulation of core regulators of cell proliferation and cell fate determinants in accordance with the progression of the germ cell cycle.</p> <p>Conclusions</p> <p>The current study provides a foundation to isolate conserved regulators of cell tip expansion and those that are unique for pollen tube growth to the female gametophyte. A transcriptomic data set is presented as a benchmark for future functional studies using developing pollen as a model. Our results demonstrated previously unknown functions of certain genes in pollen-tube tip growth. In addition, we highlighted the molecular dynamics of core cell-cycle regulators in the male gametophyte and postulated the first genetic model to account for the differential timing of spermatogenesis among angiosperms and its coordination with female gametogenesis.</p

Usability testing and evaluation of portal of the PA

Práce se zabývá testováním a hodnocením použitelnosti portálu veřejné správy. Jsou zde popsány hlavní metody testování použitelnosti a detailně rozepsáno heuristické hodnocení použitelnosti portálu veřejné správy. Pomocí navrženého postupu je realizováno heuristické hodnocení informačního systému a podle výsledků jsou popsány chyby v použitelnosti Portálu a jejich náprava.The work deals with the testing and evaluation of the usability of Portal of the Public Administration. The work describes the main methods of usability testing and details specify the heuristic evaluation of the usability of Portal of the Public Administration. Using the proposed procedure is performed heuristic evaluation of the information system and in according to the results is described errors in usability of Portal and their repair.Ústav systémového inženýrství a informatikyStudentka představila svou práci, především pak testování použitelnosti portálu veřejné správy pomocí heuristického hodnocení. Dále pak zhodnocení portálu veřejné správy a návrh použití zásad již při návrhu webu. Otázky se týkaly navržených typů heuristik a metody a jejich fáze testování. Dále pak proč studentka vyřadila některé heuristiky týkající se zdrojového kódu.Dokončená práce s úspěšnou obhajobo

Additional file 3: Table S1. of Quantitative proteomics of the tobacco pollen tube secretome identifies novel pollen tube guidance proteins important for fertilization

List of protein accessions identified from semi-in vivo pollen tube secretome and semi-in vivo total proteomes by gel-free LC/MS/MS. (XLSX 53714 kb

Additional file 1: Figure S1. of Quantitative proteomics of the tobacco pollen tube secretome identifies novel pollen tube guidance proteins important for fertilization

Semi-in vivo pollen tube secretome (SIV-PS) approach for identification and quantification of pollen tube-secreted proteins. a An improvised SIV-PS technique setup from in planta (steps 1–3) to in vitro incubation of the pollen tubes (step 4–5). The inset shows emerging pollen tubes from excised pistils. Scale bars = 2 mm. b Schematic representation of the SIV-PS workflow. c Micrographs of SIV pollen tubes showing normal pollen tube growth in bright field with streaming organelles (top panel), sperm cell formation (second panel), callose deposition and callose plugs (asterisk, third panel), and pollen tube viability assessed by Alexander stain (bottom panel). sn sperm cell nucleus, VN vegetative cell nucleus. Scale bar = 40 μM. d A tobacco pollinated pistil showing the site of stylar excision and the presumed peptide signaling flow from male and female gametophytes. (TIF 714 kb

Additional file 17: Table S6. of Quantitative proteomics of the tobacco pollen tube secretome identifies novel pollen tube guidance proteins important for fertilization

GO terms enriched in the semi-in vivo pollen tube secretome. (XLSX 472 kb