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During evolution, genomes expanded via whole-genome, segmental, tandem, and individual-gene duplications, and the emerged redundant paralogs would be eliminated or retained owing to selective neutrality or adaptive benefit and further functional divergence. Here we show that tandem paralogs can contribute adaptive quantitative benefit and thus have been retained in a lineage-specific manner. In Brassicaceae, a tandem oleosin gene cluster of five to nine paralogs encodes ample tapetum-specific oleosins located in abundant organelles called tapetosomes in flower anthers. Tapetosomes coordinate the storage of lipids and flavonoids and their transport to the adjacent maturing pollen as the coat to serve various functions. Transfer-DNA and siRNA mutants of Arabidopsis thaliana with knockout and knockdown of different tandem oleosin paralogs had quantitative and correlated loss of organized structures of the tapetosomes, pollen-coat materials, and pollen tolerance to dehydration. Complementation with the knockout paralog restored the losses. Cleomaceae is the family closest to Brassicaceae. Cleome species did not contain the tandem oleosin gene cluster, tapetum oleosin transcripts, tapetosomes, or pollen tolerant to dehydration. Cleome hassleriana transformed with an Arabidopsis oleosin gene for tapetum expression possessed primitive tapetosomes and pollen tolerant to dehydration. We propose that during early evolution of Brassicaceae, a duplicate oleosin gene mutated from expression in seed to the tapetum. The tapetum oleosin generated primitive tapetosomes that organized stored lipids and flavonoids for their effective transfer to the pollen surface for greater pollen vitality. The resulting adaptive benefit led to retention of tandem-duplicated oleosin genes for production of more oleosin and modern tapetosomes.

Extensins (EXTs), a class of hydroxyproline-rich glycoprotein with multiple Ser-Pro3–5 motifs, are known to play roles in cell wall reinforcement and environmental responses. EXTs with repetitive Tyr-X-Tyr (YXY) motifs for crosslinking are referred as crosslinking EXTs. Our comprehensive study spanned 194 algal and plant species, categorizing EXTs into seven subfamilies: classical extensins (EXT I and II), arabinogalactan-protein extensins (AGP-EXTs), proline-rich extensin-like receptor kinases (PERKs), leucine-rich repeat extensins (LRX I and II), formin homology (FH) domain-containing extensins (FH-EXTs), proline-rich, arabinogalactan proteins, conserved cysteines (PAC) domain-containing extensins (PAC I and II), and eight-cysteine motif (8CM)-containing extensins (8CM-EXTs). In the examined dataset, EXTs were detected ubiquitously in plants but infrequently in algae, except for one Coccomyxa and four Chlamydomonadales species. No crosslinking EXTs were found in Poales or certain Zingiberales species. Notably, the previously uncharacterized EXT II, PAC II, and liverwort-specific 8CM-EXTs were found to be crosslinking EXTs. EXT II, featuring repetitive YY motifs instead of the conventional YXY motif, was exclusively identified in Solanaceae. Furthermore, tandem genes encoding distinctive 8CM-EXTs specifically expressed in the germinating spores of Marchantia polymorpha. This updated classification of EXT types allows us to propose a plausible evolutionary history of EXT genes during the course of plant evolution. [ABSTRACT FROM AUTHOR]

Thraustochytrids are heterotrophic fungus-like protists that can dissolve organic matters with enzymes. Four strains, AP45, ASP1, ASP2, and ASP4, were isolated from the coastal water of Taiwan, and respectively identified as Aurantiochytrium sp., Schizochytrium sp., Parietichytrium sp., and Botryochytrium sp. based on 18S rRNA sequences. Transcriptome datasets of these four strains at days 3-5 were generated using Next Generation Sequencing technology, and screened for enzymes with potential industrial applications. Functional annotations based on KEGG database suggest that many unigenes of all four strains were related to the pathways of industrial enzymes. Most of all four strains contained homologous genes for 15 out of the 17 targeted enzymes, and had extra- and/or intra-cellular enzymatic activities, including urease, asparaginase, lipase, glucosidase, alkaline phosphatase and protease. Complete amino sequences of the first-time identified L-asparaginase and phytase in thraustochytrids were retrieved, and respectively categorized to the Type I and BPPhy families based on phylogenetic relationships, protein structural modeling and active sites. Milligram quantities of highly purified, soluble protein of urease and L-asparaginase were successfully harvested and analyzed for recombinant enzymatic activities. These analytical results highlight the diverse enzymes for wide-range applications in thraustochytrids. [ABSTRACT FROM AUTHOR]

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Lipid transfer proteins (LTPs) are small secretory proteins in plants with defined lipid-binding structures for possible lipid exocytosis. Special groups of LTPs unique to the anther tapetum are abundant, but their functions are unclear. We studied a special group of LTPs, type III LTPs, in Arabidopsis (Arabidopsis thaliana). Their transcripts were restricted to the anther tapetum, with levels peaking at the developmental stage of maximal pollen-wall exine synthesis. We constructed an LTP-Green Fluorescent Protein {LTP-GFP) plasmid, transformed it into wild-type plants, and monitored LTP-GFP in developing anthers with confocal laser scanning microscopy. LTP-GFP appeared in the tapetum and was secreted via the endoplasmic reticulum-trans-Golgi network machinery into the locule. It then moved to the microspore surface and remained as a component of exine. Immuno-transmission electron microscopy of native LTP in anthers confirmed the LTP-GFP observations. The in vivo association of LTP-GFP and exine in anthers was not observed with non-type III or structurally modified type III LTPs or in transformed exine-defective mutant plants. RNA interference knockdown of individual type III LTPs produced no observable mutant phenotypes. RNA interference knockdown of two type HI LTPs produced microscopy-observable morphologic changes in the intine underneath the exine (presumably as a consequence of changes in the exine not observed by transmission electron microscopy) and pollen susceptible to dehydration damage. Overall, we reveal a novel transfer pathway of LTPs in which LTPs bound or nonbound to exine precursors are secreted from the tapetum to become microspore exine constituents; this pathway explains the need for plentiful LTPs to incorporate into the abundant exine. [ABSTRACT FROM AUTHOR]

Lipid transfer proteins (LTPs) are small secretory proteins in plants with defined lipid-binding structures for possible lipid exocytosis. Special groups of LTPs unique to the anther tapetum are abundant, but their functions are unclear. We studied a special group of LTPs, type III LTPs, in Arabidopsis (Arabidopsis thaliana). Their transcripts were restricted to the anther tapetum, with levels peaking at the developmental stage of maximal pollen-wall exine synthesis. We constructed an LTP-Green Fluorescent Protein {LTP-GFP) plasmid, transformed it into wild-type plants, and monitored LTP-GFP in developing anthers with confocal laser scanning microscopy. LTP-GFP appeared in the tapetum and was secreted via the endoplasmic reticulum-trans-Golgi network machinery into the locule. It then moved to the microspore surface and remained as a component of exine. Immuno-transmission electron microscopy of native LTP in anthers confirmed the LTP-GFP observations. The in vivo association of LTP-GFP and exine in anthers was not observed with non-type III or structurally modified type III LTPs or in transformed exine-defective mutant plants. RNA interference knockdown of individual type III LTPs produced no observable mutant phenotypes. RNA interference knockdown of two type HI LTPs produced microscopy-observable morphologic changes in the intine underneath the exine (presumably as a consequence of changes in the exine not observed by transmission electron microscopy) and pollen susceptible to dehydration damage. Overall, we reveal a novel transfer pathway of LTPs in which LTPs bound or nonbound to exine precursors are secreted from the tapetum to become microspore exine constituents; this pathway explains the need for plentiful LTPs to incorporate into the abundant exine. [ABSTRACT FROM AUTHOR]

Abstract: The abscisic acid (ABA) hormone plays a major role in various aspects of plant growth and development, including seed maturation and germination, as well as adaptation to abiotic environmental stresses. With the purpose to identify novel ABA-regulated genes, we tried to genome-wide identification of ABA-related genes using computational approach by searching ABA-responsive elements (ABREs, T/CACGTGT/GC) in entire Arabidopsis genome. 137 ABA-regulated candidate genes were found to possess two or more ABREs in the 1200-bp upstream region of the translation start site. In an attempt to monitor the gene expression of these candidates in response to ABA, 16 genes were selected and examined by RT-PCR. Of these 16, seven are unclassified genes in Arabidopsis, 12 candidates were up-regulated by ABA and four had equal expression levels as control. In addition, Northern blot analysis indicated that some candidates were further shown to differentially respond to various stresses. The results suggested that these ABA-regulated genes may consist of other cis-acting elements responsive to abiotic stresses and confirmed that in silico cloning of ABA-regulated genes through Arabidopsis genomic sequence database mining is a fast and reliable approach. [Copyright &y& Elsevier]

Image stacking is a crucial method for micro or macro photography. It captures images at different focal planes and then merges them into a single, all-in-focus image with extended focus. This method has been extensively used for digital documentation by scientists working at museums or research institutions. However, the traditional image stacking method relies on expensive instruments to conduct precise image stacking using a computer-based stepper motor controller. In this study, we reported how to conduct image focus extensions with comparable quality to those done by a motorized stepper using a cost-effective instrument setting and an efficient manual stacking method. This method provides a shorter operation time and capability to capture images of living objects and high flexibility in obtaining the images of objects from cm to mm scale. However, it also has some limitations, including the inability to control aperture and exposure time, relatively short working distance at high magnification, requires additional steps to convert the video into images, and heavily relies on the user's manual observation prior to a video recording. Nevertheless, the authors believe that the current method can be applied as an alternative method to conduct image stacking. The development of such an instrument and method offers a promising avenue for scientists to perform image stacking with greater flexibility and speed in macro photography. [ABSTRACT FROM AUTHOR]

One ubiquinone derivative, cinnamomeaol acid (1), was isolated from the mycelia of Antrodia cinnamomea BCRC 36799 by solid-state fermentation with adlay. The structure was established by spectroscopic analyses including 1D and 2D NMR (1H, 13C, COSY, HSQC, HMBC, and NOESY) as well as HR-ESI-MS. [ABSTRACT FROM AUTHOR]