Cytokinin activity - transport and homeostasis at the whole plant, cell, and subcellular levels.

Cytokinins (CKs) are important plant hormones that regulate a variety of biological processes implicated in plant development and stress responses. Here, we summarize the most recent advances in discovering and characterizing the membrane transporters involved in long- and short-distance translocation of CKs and their significance in CK signal activity. We highlight the discovery of PUP7 and PUP21 tonoplast-localized transporters and propose potential mechanisms for CK subcellular homeostasis. Finally, we discuss the importance of subcellular hormone transport in light of the localization of histidine kinase receptors of CKs at the endoplasmic reticulum and plasma membrane.

Multi-Knock-a multi-targeted genome-scale CRISPR toolbox to overcome functional redundancy in plants.

Plant genomes are characterized by large and complex gene families that often result in similar and partially overlapping functions. This genetic redundancy severely hampers current efforts to uncover novel phenotypes, delaying basic genetic research and breeding programmes. Here we describe the development and validation of Multi-Knock, a genome-scale clustered regularly interspaced short palindromic repeat toolbox that overcomes functional redundancy in Arabidopsis by simultaneously targeting multiple gene-family members, thus identifying genetically hidden components. We computationally designed 59,129 optimal single-guide RNAs that each target two to ten genes within a family at once. Furthermore, partitioning the library into ten sublibraries directed towards a different functional group allows flexible and targeted genetic screens. From the 5,635 single-guide RNAs targeting the plant transportome, we generated over 3,500 independent Arabidopsis lines that allowed us to identify and characterize the first known cytokinin tonoplast-localized transporters in plants. With the ability to overcome functional redundancy in plants at the genome-scale level, the developed strategy can be readily deployed by scientists and breeders for basic research and to expedite breeding efforts.

[Spawning substrates of cuttlefish: Type, function, and application]. / 乌贼类产卵附着基的类型、功能和应用.

The fertilized eggs of cuttlefish are sticky eggs. Cuttlefish parents prefer to lay eggs on the attached substrates, which help increase the number of eggs and the hatching rate of fertilized eggs. If egg-attached substrates are sufficient, cuttlefish spawning will be reduced or even delayed. With the advances in the construction of marine nature reserves and research on artificial enrichment techniques, domestic and international experts have conducted research on different types and configurations of attachment substrates around cuttlefish resource enhancement. Based on the source of the substrates, we classified cuttlefish spawning substrates into two types, natural and artificial. By summarizing the differences, advantages, and disadvantages of the common economic cuttlefish spawning substrates in offshore areas worldwide, we sort out the functions of two different types of attachment bases, and discussed the practical applications of natural and artificial egg-attached substrates in spawning ground restoration and artificial enrichment. We proposed several thoughts on the future research directions of cuttlefish spawning attachment substrates, aiming to provide reasonable suggestions for cuttlefish habitat restoration, cuttlefish breeding and sustainable development of fishery resources.

ABCB-mediated shootward auxin transport feeds into the root clock.

Although strongly influenced by environmental conditions, lateral root (LR) positioning along the primary root appears to follow obediently an internal spacing mechanism dictated by auxin oscillations that prepattern the primary root, referred to as the root clock. Surprisingly, none of the hitherto characterized PIN- and ABCB-type auxin transporters seem to be involved in this LR prepatterning mechanism. Here, we characterize ABCB15, 16, 17, 18, and 22 (ABCB15-22) as novel auxin-transporting ABCBs. Knock-down and genome editing of this genetically linked group of ABCBs caused strongly reduced LR densities. These phenotypes were correlated with reduced amplitude, but not reduced frequency of the root clock oscillation. High-resolution auxin transport assays and tissue-specific silencing revealed contributions of ABCB15-22 to shootward auxin transport in the lateral root cap (LRC) and epidermis, thereby explaining the reduced auxin oscillation. Jointly, these data support a model in which LRC-derived auxin contributes to the root clock amplitude.

The GORKY glycoalkaloid transporter is indispensable for preventing tomato bitterness.

Fruit taste is determined by sugars, acids and in some species, bitter chemicals. Attraction of seed-dispersing organisms in nature and breeding for consumer preferences requires reduced fruit bitterness. A key metabolic shift during ripening prevents tomato fruit bitterness by eliminating α-tomatine, a renowned defence-associated Solanum alkaloid. Here, we combined fine mapping with information from 150 resequenced genomes and genotyping a 650-tomato core collection to identify nine bitter-tasting accessions including the 'high tomatine' Peruvian landraces reported in the literature. These 'bitter' accessions contain a deletion in GORKY, a nitrate/peptide family transporter mediating α-tomatine subcellular localization during fruit ripening. GORKY exports α-tomatine and its derivatives from the vacuole to the cytosol and this facilitates the conversion of the entire α-tomatine pool to non-bitter forms, rendering the fruit palatable. Hence, GORKY activity was a notable innovation in the process of tomato fruit domestication and breeding.

Cell kinetics of auxin transport and activity in Arabidopsis root growth and skewing.

Auxin is a key regulator of plant growth and development. Local auxin biosynthesis and intercellular transport generates regional gradients in the root that are instructive for processes such as specification of developmental zones that maintain root growth and tropic responses. Here we present a toolbox to study auxin-mediated root development that features: (i) the ability to control auxin synthesis with high spatio-temporal resolution and (ii) single-cell nucleus tracking and morphokinetic analysis infrastructure. Integration of these two features enables cutting-edge analysis of root development at single-cell resolution based on morphokinetic parameters under normal growth conditions and during cell-type-specific induction of auxin biosynthesis. We show directional auxin flow in the root and refine the contributions of key players in this process. In addition, we determine the quantitative kinetics of Arabidopsis root meristem skewing, which depends on local auxin gradients but does not require PIN2 and AUX1 auxin transporter activities. Beyond the mechanistic insights into root development, the tools developed here will enable biologists to study kinetics and morphology of various critical processes at the single cell-level in whole organisms.

Cell-type action specificity of auxin on Arabidopsis root growth.

The plant hormone auxin plays a critical role in root growth and development; however, the contributions or specific roles of cell-type auxin signals in root growth and development are not well understood. Here, we mapped tissue and cell types that are important for auxin-mediated root growth and development by manipulating the local response and synthesis of auxin. Repressing auxin signaling in the epidermis, cortex, endodermis, pericycle or stele strongly inhibited root growth, with the largest effect observed in the endodermis. Enhancing auxin signaling in the epidermis, cortex, endodermis, pericycle or stele also caused reduced root growth, albeit to a lesser extent. Moreover, we established that root growth was inhibited by enhancement of auxin synthesis in specific cell types of the epidermis, cortex and endodermis, whereas increased auxin synthesis in the pericycle and stele had only minor effects on root growth. Our study thus establishes an association between cellular identity and cell type-specific auxin signaling that guides root growth and development.

Autophagy-Related Beclin 1 and Head and Neck Cancers.

Beclin 1, a positive regulator of autophagy, behaves as a double-edged sword in tumorigenesis. Beclin 1 contributes to tumor suppression by removing defective or damaged organelles and other cellular components; however, its activity can also stimulate cancer initiation and progression. In head and neck cancer, Beclin 1 overexpression promotes autophagy, which limits DNA damage and chromosomal instability and increases necrosis and inflammation by impacting apoptotic and autophagic pathways. This paper reviews the relationship between Beclin 1, carcinogenesis and head and neck cancer prognosis.

Transcriptional Basis for Differential Thermosensitivity of Seedlings of Various Tomato Genotypes.

Transcriptional reprograming after the exposure of plants to elevated temperatures is a hallmark of stress response which is required for the manifestation of thermotolerance. Central transcription factors regulate the stress survival and recovery mechanisms and many of the core responses controlled by these factors are well described. In turn, pathways and specific genes contributing to variations in the thermotolerance capacity even among closely related plant genotypes are not well defined. A seedling-based assay was developed to directly compare the growth and transcriptome response to heat stress in four tomato genotypes with contrasting thermotolerance. The conserved and the genotype-specific alterations of mRNA abundance in response to heat stress were monitored after exposure to three different temperatures. The transcripts of the majority of genes behave similarly in all genotypes, including the majority of heat stress transcription factors and heat shock proteins, but also genes involved in photosynthesis and mitochondrial ATP production. In turn, genes involved in hormone and RNA-based regulation, such as auxin- and ethylene-related genes, or transcription factors like HsfA6b, show a differential regulation that associates with the thermotolerance pattern. Our results provide an inventory of genes likely involved in core and genotype-dependent heat stress response mechanisms with putative role in thermotolerance in tomato seedlings.

Natural variation in HsfA2 pre-mRNA splicing is associated with changes in thermotolerance during tomato domestication.

Wild relatives of crops thrive in habitats where environmental conditions can be restrictive for productivity and survival of cultivated species. The genetic basis of this variability, particularly for tolerance to high temperatures, is not well understood. We examined the capacity of wild and cultivated accessions to acclimate to rapid temperature elevations that cause heat stress (HS). We investigated genotypic variation in thermotolerance of seedlings of wild and cultivated accessions. The contribution of polymorphisms associated with thermotolerance variation was examined regarding alterations in function of the identified gene. We show that tomato germplasm underwent a progressive loss of acclimation to strong temperature elevations. Sensitivity is associated with intronic polymorphisms in the HS transcription factor HsfA2 which affect the splicing efficiency of its pre-mRNA. Intron splicing in wild species results in increased synthesis of isoform HsfA2-II, implicated in the early stress response, at the expense of HsfA2-I which is involved in establishing short-term acclimation and thermotolerance. We propose that the selection for modern HsfA2 haplotypes reduced the ability of cultivated tomatoes to rapidly acclimate to temperature elevations, but enhanced their short-term acclimation capacity. Hence, we provide evidence that alternative splicing has a central role in the definition of plant fitness plasticity to stressful conditions.

The repressor and co-activator HsfB1 regulates the major heat stress transcription factors in tomato.

Plants code for a multitude of heat stress transcription factors (Hsfs). Three of them act as central regulators of heat stress (HS) response in tomato (Solanum lycopersicum). HsfA1a regulates the initial response, and HsfA2 controls acquired thermotolerance. HsfB1 is a transcriptional repressor but can also act as co-activator of HsfA1a. Currently, the mode of action and the relevance of the dual function of HsfB1 remain elusive. We examined this in HsfB1 overexpression or suppression transgenic tomato lines. Proteome analysis revealed that HsfB1 overexpression stimulates the co-activator function of HsfB1 and consequently the accumulation of HS-related proteins under non-stress conditions. Plants with enhanced levels of HsfB1 show aberrant growth and development but enhanced thermotolerance. HsfB1 suppression has no significant effect prior to stress. Upon HS, HsfB1 suppression strongly enhances the induction of heat shock proteins due to the higher activity of other HS-induced Hsfs, resulting in increased thermotolerance compared with wild-type. Thereby, HsfB1 acts as co-activator of HsfA1a for several Hsps, but as a transcriptional repressor on other Hsfs, including HsfA1b and HsfA2. The dual function explains the activation of chaperones to enhance protection and regulate the balance between growth and stress response upon deviations from the homeostatic levels of HsfB1.

Synthesis and luminescence properties of brick-shaped lanthanum-organic frameworks with mesoporous and macroporous architectures.

Generally, metal-organic frameworks (MOFs) are made up from kinds of repeating microporous structure. Here, a series of Eu3+ ions activated terephthalate-based lanthanum-organic frameworks (La-MOFs) was synthesized by a hydrothermal reaction. By controlling the reaction time, we obtained some unique brick-shaped La-MOFs in a micron scale size range, and these La-MOFs showed tunable mesoporous and macroporous architectures. It is speculated that the change in the composition and structure of building units results in the formation of this mesoporous and macroporous heterogeneous architectures. Powder X-ray diffraction patterns and Eu3+ luminescence behavior support the speculation.

Alternative splicing in tomato pollen in response to heat stress.

Alternative splicing (AS) is a key control mechanism influencing signal response cascades in different developmental stages and under stress conditions. In this study, we examined heat stress (HS)-induced AS in the heat sensitive pollen tissue of two tomato cultivars. To obtain the entire spectrum of HS-related AS, samples taken directly after HS and after recovery were combined and analysed by RNA-seq. For nearly 9,200 genes per cultivar, we observed at least one AS event under HS. In comparison to control, for one cultivar we observed 76% more genes with intron retention (IR) or exon skipping (ES) under HS. Furthermore, 2,343 genes had at least one transcript with IR or ES accumulated under HS in both cultivars. These genes are involved in biological processes like protein folding, gene expression and heat response. Transcriptome assembly of these genes revealed that most of the alternative spliced transcripts possess truncated coding sequences resulting in partial or total loss of functional domains. Moreover, 141 HS specific and 22 HS repressed transcripts were identified. Further on, we propose AS as layer of stress response regulating constitutively expressed genes under HS by isoform abundance.

Unfolded protein response in pollen development and heat stress tolerance.

KEY MESSAGE: Importance of the UPR for pollen. Pollen is particularly sensitive to environmental conditions that disturb protein homeostasis, such as higher temperatures. Their survival is dependent on subcellular stress response systems, one of which maintains protein homeostasis in the endoplasmic reticulum (ER). Disturbance of ER proteostasis due to stress leads to the activation of the unfolded protein response (UPR) that mitigates stress damage mainly by increasing ER-folding capacity and reducing folding demands. The UPR is controlled by ER membrane-associated transcription factors and an RNA splicing factor. They are important components of abiotic stress responses including general heat stress response and thermotolerance. In addition to responding to environmental stresses, the UPR is implicated in developmental processes required for successful male gametophyte development and fertilization. Consequently, defects in the UPR can lead to pollen abortion and male sterility. Several UPR components are involved in the elaboration of the ER network, which is required for pollen germination and polar tube growth. Transcriptome and proteome analyses have shown that components of the ER-folding machinery and the UPR are upregulated at specific stages of pollen development supporting elevated demands for secretion. Furthermore, genetic studies have revealed that knockout mutants of UPR genes are defective in producing viable or competitive pollen. In this review, we discuss recent findings regarding the importance of the UPR for both pollen development and stress response.

HsfA2 Controls the Activity of Developmentally and Stress-Regulated Heat Stress Protection Mechanisms in Tomato Male Reproductive Tissues.

Male reproductive tissues are more sensitive to heat stress (HS) compared to vegetative tissues, but the basis of this phenomenon is poorly understood. Heat stress transcription factors (Hsfs) regulate the transcriptional changes required for protection from HS In tomato (Solanum lycopersicum), HsfA2 acts as coactivator of HsfA1a and is one of the major Hsfs accumulating in response to elevated temperatures. The contribution of HsfA2 in heat stress response (HSR) and thermotolerance was investigated in different tissues of transgenic tomato plants with suppressed HsfA2 levels (A2AS). Global transcriptome analysis and immunodetection of two major Hsps in vegetative and reproductive tissues showed that HsfA2 regulates subsets of HS-induced genes in a tissue-specific manner. Accumulation of HsfA2 by a moderate HS treatment enhances the capacity of seedlings to cope with a subsequent severe HS, suggesting an important role for HsfA2 in regulating acquired thermotolerance. In pollen, HsfA2 is an important coactivator of HsfA1a during HSR HsfA2 suppression reduces the viability and germination rate of pollen that received the stress during the stages of meiosis and microspore formation but had no effect on more advanced stages. In general, pollen meiocytes and microspores are characterized by increased susceptibility to HS due to their lower capacity to induce a strong HSR This sensitivity is partially mitigated by the developmentally regulated expression of HsfA2 and several HS-responsive genes mediated by HsfA1a under nonstress conditions. Thereby, HsfA2 is an important factor for the priming process that sustains pollen thermotolerance during microsporogenesis.