Fertilisation and cell cycle in angiosperms

Fertilisation is the result of successful fusion of female and male gametes, forming a zygote that develops into the embryo. From the perspective of cell cycle control, fertilisation is a hat‐trick. Firstly, the generation of gametes depends on the generation of haploid spores by meiosis and a sequence of mitotic divisions. Mitosis is required to produce the cells of the gametophyte, closely associated with differential fate acquisition. Secondly, cell cycle progression in both male and female gametes has to be synchronised in order to avoid chromosomal imbalance at karyogamy, and last but not least, the cell cycle should only be relaunched after a successful fusion. Here, we seek to survey our current knowledge of these processes from a cell cycle perspective and explore possible mechanisms involved in cell cycle control and coordination

Phosphatidic acid counteracts S-RNase signaling in pollen by stabilizing the actin cytoskeleton

S-RNase is the female determinant of self-incompatibility (SI) in pear (Pyrus bretschneideri). After translocation to the pollen tube, S-RNase degrades rRNA and induces pollen tube death in an S-haplotype-specific manner. In this study, we found that the actin cytoskeleton is a target of P. bretschneideri S-RNase (PbrS-RNase) and uncovered a mechanism that involves phosphatidic acid (PA) and protects the pollen tube from PbrS-RNase cytotoxicity. PbrS-RNase interacts directly with PbrActin1 in an S-haplotype-independent manner, causing the actin cytoskeleton to depolymerize and promoting programmed cell death in the self-incompatible pollen tube. Pro-156 of PbrS-RNase is essential for the PbrS-RNase-PbrActin1 interaction, and the actin cytoskeleton-depolymerizing function of PbrS-RNase does not require its RNase activity. PbrS-RNase cytotoxicity enhances the expression of phospholipase D (PbrPLDδ1), resulting in increased PA levels in the incompatible pollen tube. PbrPLDδ1-derived PA initially prevents depolymerization of the actin cytoskeleton elicited by PbrS-RNase and delays the SI signaling that leads to pollen tube death. This work provides insights into the orchestration of the S-RNase-based SI response, in which increased PA levels initially play a protective role in incompatible pollen, until sustained PbrS-RNase activity reaches the point of no return and pollen tube growth ceases

Phytotracker, an information management system for easy recording and tracking of plants, seeds and plasmids

Background A large number of different plant lines are produced and maintained in a typical plant research laboratory, both as seed stocks and in active growth. These collections need careful and consistent management to track and maintain them properly, and this is a particularly pressing issue in laboratories undertaking research involving genetic manipulation due to regulatory requirements. Researchers and PIs need to access these data and collections, and therefore an easy-to-use plant-oriented laboratory information management system that implements, maintains and displays the information in a simple and visual format would be of great help in both the daily work in the lab and in ensuring regulatory compliance. Results Here, we introduce ‘Phytotracker’, a laboratory management system designed specifically to organise and track plasmids, seeds and growing plants that can be used in mixed platform environments. Phytotracker is designed with simplicity of user operation and ease of installation and management as the major factor, whilst providing tracking tools that cover the full range of activities in molecular genetics labs. It utilises the cross-platform Filemaker relational database, which allows it to be run as a stand-alone or as a server-based networked solution available across all workstations in a lab that can be internet accessible if desired. It can also be readily modified or customised further. Phytotracker provides cataloguing and search functions for plasmids, seed batches, seed stocks and plants growing in pots or trays, and allows tracking of each plant from seed sowing, through harvest to the new seed batch and can print appropriate labels at each stage. The system enters seed information as it is transferred from the previous harvest data, and allows both selfing and hybridization (crossing) to be defined and tracked. Transgenic lines can be linked to their plasmid DNA source. This ease of use and flexibility helps users to reduce their time needed to organise their plants, seeds and plasmids and to maintain laboratory continuity involving multiple workers. Conclusion We have developed and used Phytotracker for over five years and have found it has been an intuitive, powerful and flexible research tool in organising our plasmid, seed and plant collections requiring minimal maintenance and training for users. It has been developed in an Arabidopsis molecular genetics environment, but can be readily adapted for almost any plant laboratory research

Mutation of Arabidopsis copper-containing amine oxidase gene AtCuAOδ alters polyamines, reduces gibberellin content and affects development

Polyamines (PAs) are essential metabolites in plants performing multiple functions during growth and development. Copper-containing amine oxidases (CuAOs) catalyse the catabolism of PAs and in Arabidopsis thaliana are encoded by a gene family. Two mutants of one gene family member, AtCuAOδ, showed delayed seed germination, leaf emergence, and flowering time. The height of the primary inflorescence shoot was reduced, and developmental leaf senescence was delayed. Siliques were significantly longer in mutant lines and contained more seeds. The phenotype of AtCuAOδ over-expressors was less affected. Before flowering, there was a significant increase in putrescine in AtCuAOδ mutant leaves compared to wild type (WT), while after flowering both spermidine and spermine concentrations were significantly higher than in WT leaves. The expression of GA (gibberellic acid) biosynthetic genes was repressed and the content of GA1, GA7, GA8, GA9, and GA20 was reduced in the mutants. The inhibitor of copper-containing amine oxidases, aminoguanidine hydrochloride, mimicked the effect of AtCuAOδ mutation on WT seed germination. Delayed germination, reduced shoot height, and delayed flowering in the mutants were rescued by GA3 treatment. These data strongly suggest AtCuAOδ is an important gene regulating PA homeostasis, and that a perturbation of PAs affects plant development through a reduction in GA biosynthesis

Expression of the Arabidopsis redox-related LEA protein, SAG21 is regulated by ERF, NAC and WRKY transcription factors

SAG21/LEA5 is an unusual late embryogenesis abundant protein in Arabidopsis thaliana, that is primarily mitochondrially located and may be important in regulating translation in both chloroplasts and mitochondria. SAG21 expression is regulated by a plethora of abiotic and biotic stresses and plant growth regulators indicating a complex regulatory network. To identify key transcription factors regulating SAG21 expression, yeast-1-hybrid screens were used to identify transcription factors that bind the 1685 bp upstream of the SAG21 translational start site. Thirty-three transcription factors from nine different families bound to the SAG21 promoter, including members of the ERF, WRKY and NAC families. Key binding sites for both NAC and WRKY transcription factors were tested through site directed mutagenesis indicating the presence of cryptic binding sites for both these transcription factor families. Co-expression in protoplasts confirmed the activation of SAG21 by WRKY63/ABO3, and SAG21 upregulation elicited by oligogalacturonide elicitors was partially dependent on WRKY63, indicating its role in SAG21 pathogen responses. SAG21 upregulation by ethylene was abolished in the erf1 mutant, while wound-induced SAG21 expression was abolished in anac71 mutants, indicating SAG21 expression can be regulated by several distinct transcription factors depending on the stress condition

Engineering of Plants to exhibit Self-Incompatibility

Self-incompatibility (Sl) of the common field poppy (Papaver rhoeas) depends on interaction of a pollen transmembrane protein with a pistil ligand protein both encoded by multi-allelic genes at the S locus. Such a locus can be used to confer SI on other plant species

Cellular mechanisms for pollen tube growth inhibition in gametophytic self-incompatibility

Self-incompatibility (SI) is a mechanism used by angiosperms to prevent self-fertilization. Here we review current knowledge of two different gametophytic SI systems at the cellular level, revealing different mechanisms that interfere with pollen tube growth. In the Solanaceae, Rosaceae, and Scrophulariaceae, SI is controlled by an interaction between a pistil component, S-RNase, and a pollen component, an F-box protein, SLF/SFB. While a variety of models focused on ubiquitylation have been explored, it is still unclear exactly how the S-RNase based system operates at the cellular level. In Papaver, entirely different S-proteins act as signalling ligands that trigger a Ca 2+-dependent signalling cascade that results in programmed cell death (PCD). Although the pollen S-receptor has not been identified in Papaver, the mechanisms involved in inhibiting incompatible pollen are better understood

Revision techniques for failed PD catheters: outcome in a University Hospital

Purpose: A disfunctioning peritoneal dialysis (PD) catheter has been reported in upto 35%. We report different salvage techniques used and its outcome. Methods: We retrospectively reviewed our database on PD patients from 2009 to 2014. Operational data and data on PD catheter function were checked. Results: From 2009 to 2014, we operated on 32 patients. Malfunction of their PD catheter was found in 23 patients and hernias in nine patients. Different laparoscopic techniques were used, including reposition of the catheter, fixation of the catheter, removal of adhesions and omentectomy. Of these 23 patients, 18 (78%) had a normal functioning catheter after the operation. Nine patients of the total of 32 patients had a correction of their hernia. In all cases, a non-absorbable mesh was used. Of those nine patients with a hernia, eight (89%) had a normal functioning catheter after the procedure. Conclusions: The salvage of the catheter by means of a laparoscopic procedure or correction of a hernia is worthwhile and can save up to 81% of the catheters