Developmental stages and episode-specific regulatory genes in andromonoecious melon flower development.

BACKGROUND AND AIMS: Given the lack of specific studies on floral development in melon (Cucumis melo L.), we carried out an extensive study involving morphological and transcriptomic analyses to characterize floral development in this species. METHODS: Using an andromonoecious line, we analysed the development of floral buds in male and hermaphrodite flowers with both light microscopy and scanning electron microscopy. Based on flower lengths, we established a correlation between the developmental stages and four main episodes of floral development and conducted an extensive RNA sequencing analysis of these episodes. KEY RESULTS: We identified 12 stages of floral development, from the appearance of the floral meristems to anthesis. The main structural differences between male and hermaphrodite flowers appeared between stages 6 and 7; later stages of development leading to the formation of organs and structures in both types of flowers were also described. We analysed the gene expression patterns of the four episodes in flower development to find the genes that were specific to each given episode. Among others, we identified genes that defined the passage from one episode to the next according to the ABCDE model of floral development. CONCLUSIONS: This work combines a detailed morphological analysis and a comprehensive transcriptomic study to enable characterization of the structural and molecular mechanisms that determine the floral development of an andromonoecious genotype in melon. Taken together, our results provide a first insight into gene regulation networks in melon floral development that are crucial for flowering and pollen formation, highlighting potential targets for genetic manipulation to improve crop yield of melon in the future.

Autofluorescence Imaging to Evaluate Red Algae Physiology.

Red algae (Rhodophyta) contain phycobiliproteins and colonize habitats with dim light, however some (e.g., some Chroothece species) can also develop in full sunshine. Most rhodophytes are red, however some can appear bluish, depending on the proportion of blue and red biliproteins (phycocyanin and phycoerythrin). Different phycobiliproteins can capture light at diverse wavelengths and transmit it to chlorophyll a, which makes photosynthesis under very different light conditions possible. These pigments respond to habitat changes in light, and their autofluorescence can help to study biological processes. Using Chroothece mobilis as a model organism and the spectral lambda scan mode in a confocal microscope, the adaptation of photosynthetic pigments to different monochromatic lights was studied at the cellular level to guess the species' optimal growth conditions. The results showed that, even when the studied strain was isolated from a cave, it adapted to both dim and medium light intensities. The presented method is especially useful for studying photosynthetic organisms that do not grow or grow very slowly under laboratory conditions, which is usually the case for those living in extreme habitats.

Deciphering the Role and Signaling Pathways of PKCα in Luminal A Breast Cancer Cells.

Protein kinase C (PKC) comprises a family of highly related serine/threonine protein kinases involved in multiple signaling pathways, which control cell proliferation, survival, and differentiation. The role of PKCα in cancer has been studied for many years. However, it has been impossible to establish whether PKCα acts as an oncogene or a tumor suppressor. Here, we analyzed the importance of PKCα in cellular processes such as proliferation, migration, or apoptosis by inhibiting its gene expression in a luminal A breast cancer cell line (MCF-7). Differential expression analysis and phospho-kinase arrays of PKCα-KD vs. PKCα-WT MCF-7 cells identified an essential set of proteins and oncogenic kinases of the JAK/STAT and PI3K/AKT pathways that were down-regulated, whereas IGF1R, ERK1/2, and p53 were up-regulated. In addition, unexpected genes related to the interferon pathway appeared down-regulated, while PLC, ERBB4, or PDGFA displayed up-regulated. The integration of this information clearly showed us the usefulness of inhibiting a multifunctional kinase-like PKCα in the first step to control the tumor phenotype. Then allowing us to design a possible selection of specific inhibitors for the unexpected up-regulated pathways to further provide a second step of treatment to inhibit the proliferation and migration of MCF-7 cells. The results of this study suggest that PKCα plays an oncogenic role in this type of breast cancer model. In addition, it reveals the signaling mode of PKCα at both gene expression and kinase activation. In this way, a wide range of proteins can implement a new strategy to fine-tune the control of crucial functions in these cells and pave the way for designing targeted cancer therapies.

Confocal Microscopy in Ecophysiological Studies of Algae: A Door to Understanding Autofluorescence in Red Algae.

Alga in the genus Chroothece have been reported mostly from aquatic or subaerial continental environments, where they grow in extreme conditions. The strain Chroothece mobilis MAESE 20.29 was exposed to different light intensities, red and green monochromatic light, ultraviolet (UV) radiation, high nitrogen concentrations, and high salinity to assess the effect of those environmental parameters on its growth. Confocal laser scanning microscopy (CLSM) was used as an "in vivo" noninvasive single-cell method for the study. The strain seemed to prefer fairly high light intensities and showed a significant increase in allophycocyanin (APC) and chlorophyll a [photosystem I (PSI) and photosystem II (PSII)] fluorescence with 330 and 789 µM/cm2/s intensities. Green monochromatic light promoted a significant increase in the fluorescence of APC and chlorophyll a (PSI and PSII). UV-A significantly decreased phycocyanin and increased APC, while UV-A + B showed a greater decreasing effect on c-Phycocyanin but did not significantly change concentrations of APC. The increase in nitrogen concentration in the culture medium significantly and negatively affected all pigments, and no effect was observed with an increase in salinity. Our data show that CLSM represents a very powerful tool for ecological research of microalgae in small volumes and may contribute to the knowledge of phycobiliproteins in vivo behavior and the parameters for the large-scale production of these pigments.

Structural characterization of the Rabphilin-3A-SNAP25 interaction.

Membrane fusion is essential in a myriad of eukaryotic cell biological processes, including the synaptic transmission. Rabphilin-3A is a membrane trafficking protein involved in the calcium-dependent regulation of secretory vesicle exocytosis in neurons and neuroendocrine cells, but the underlying mechanism remains poorly understood. Here, we report the crystal structures and biochemical analyses of Rabphilin-3A C2B-SNAP25 and C2B-phosphatidylinositol 4,5-bisphosphate (PIP2) complexes, revealing how Rabphilin-3A C2 domains operate in cooperation with PIP2/Ca2+ and SNAP25 to bind the plasma membrane, adopting a conformation compatible to interact with the complete SNARE complex. Comparisons with the synaptotagmin1-SNARE show that both proteins contact the same SNAP25 surface, but Rabphilin-3A uses a unique structural element. Data obtained here suggest a model to explain the Ca2+-dependent fusion process by membrane bending with a myriad of variations depending on the properties of the C2 domain-bearing protein, shedding light to understand the fine-tuning control of the different vesicle fusion events.