A Novel R2R3-MYB Gene LoMYB33 From Lily Is Specifically Expressed in Anthers and Plays a Role in Pollen Development

Lily (Lilium spp.) is an important commercial flower crop, but its market popularity and applications are adversely affected by severe pollen pollution. Many studies have examined pollen development in model plants, but few studies have been conducted on flower crops such as lily. GAMYBs are a class of R2R3-MYB transcription factors and play important roles in plant development and biotic resistance; their functions vary in different pathways, and many of them are involved in anther development. However, their function and regulatory role in lily remain unclear. Here, the GAMYB homolog LoMYB33 was isolated and identified from lily. The open reading frame of LoMYB33 was 1620 bp and encoded a protein with 539 amino acids localized in the nucleus and cytoplasm. Protein sequence alignment showed that LoMYB33 contained a conserved R2R3 domain and three BOX motifs (BOX1, BOX2, and BOX3), which were unique to the GAMYB family. LoMYB33 had transcriptional activation activity, and its transactivation domain was located within 90 amino acids of the C-terminal. LoMYB33 was highly expressed during the late stages of anther development, especially in pollen. Analysis of the promoter activity of LoMYB33 in transgenic Arabidopsis revealed that the LoMYB33 promoter was highly activated in the pollen of stage 12 to 13 flowers. Overexpression of LoMYB33 in Arabidopsis significantly retarded growth; the excess accumulation of LoMYB33 also negatively affected normal anther development, which generated fewer pollen grains and resulted in partial male sterility in transgenic plants. Silencing of LoMYB33 in lily also greatly decreased the amount of pollen. Overall, our results suggested that LoMYB33 might play an important role in the anther development and pollen formation of lily.

Full-length transcriptome analysis provides new insights into the early bolting occurrence in medicinal Angelica sinensis

Angelica sinensis (Oliv.) Diels root part is an integral component of traditional Chinese medicine, widely prescribed to improve blood circulation and blood stasis. However, early bolting of A. sinensis compromises the quality of the roots and hence is a major limitation for yield of medicinal materials. To date, little information about the molecular mechanisms underlying bolting is available for this important medicinal plant. To identify genes putatively involved in early bolting, we have conducted the transcriptome analysis of the shoot tips of the early-bolting plants and non-bolting (normal) plants of A. sinensis, respectively, using a combination of third-generation sequencing and next-generation sequencing. A total of 43,438 non-redundant transcripts were collected and 475 unique differentially expressed genes (DEGs) were identified. Gene annotation and functional analyses revealed that DEGs were highly involved in plant hormone signaling and biosynthesis pathways, three main flowering pathways, pollen formation, and very-long-chain fatty acids biosynthesis pathways. The levels of endogenous hormones were also changed significantly in the early bolting stage of A. sinensis. This study provided new insights into the transcriptomic control of early bolting in A. sinensis, which could be further applied to enhance the yield of medicinally important raw materials.

Microsporogenesis and pollen formation in Larch Sukachev (Larix Sukaczewii Djil.) on permanent forest seed plot in Semiluksky forestry

The formation of male and female generative buds in the Sukachev larch in the conditions of Voronezh in the second decade of August is considered. The behavior of chromosomes in the meta-, ana -, and telophase of the first and second divisions of meiosis is analyzed. In each phase, 150–200 microsporocytes were taken into account. According to the results of the conducted studies, it was revealed that the microsporogenesis of larch proceeded asynchronously. A significant proportion of the disorders are caused by chromosome lag and the formation of bridges, the formation of a hexad, and the release of chromosomes outside the division spindle. The viability of Sukachev larch pollen is estimated to be high. A small number of disturbances in the process of meiotic divisions and the formation of gametophytes did not lead to the formation of a significant amount of sterile pollen. The average pollen size varies between 82.18–86.4 microns. Pollen has a spherical shape

A MADS-Box Gene CiMADS43 Is Involved in Citrus Flowering and Leaf Development through Interaction with CiAGL9

MADS-box genes are involved in various developmental processes including vegetative development, flower architecture, flowering, pollen formation, seed and fruit development. However, the function of most MADS-box genes and their regulation mechanism are still unclear in woody plants compared with model plants. In this study, a MADS-box gene (CiMADS43) was identified in citrus. Phylogenetic and sequence analysis showed that CiMADS43 is a GOA-like Bsister MADS-box gene. It was localized in the nucleus and as a transcriptional activator. Overexpression of CiMADS43 promoted early flowering and leaves curling in transgenic Arabidopsis. Besides, overexpression or knockout of CiMADS43 also showed leaf curl phenotype in citrus similar to that of CiMADS43 overexpressed in Arabidopsis. Protein–protein interaction found that a SEPALLATA (SEP)-like protein (CiAGL9) interacted with CiMADS43 protein. Interestingly, CiAGL9 also can bind to the CiMADS43 promoter and promote its transcription. Expression analysis also showed that these two genes were closely related to seasonal flowering and the development of the leaf in citrus. Our findings revealed the multifunctional roles of CiMADS43 in the vegetative and reproductive development of citrus. These results will facilitate our understanding of the evolution and molecular mechanisms of MADS-box genes in citrus.

Pollination.

Pollination of flowers is the transfer of pollen grains (haploid male spores) from the anther (part of the androecium) to the stigma (part of the gynoecium) by biotic or abiotic factors (Sliwinska and Bewley, 2014). For seed and fruit production of agricultural crops the main pollinating agents are wind and insects (George, 2011). After a pollen grain is transferred to a receptive stigma, it absorbs water from the stigma surface and germinates. A pollen tube then grows down into the stigma, through the gynoecium and through the apical micropyle; from there it grows into an ovule in the ovary and double fertilisation then takes place. Two sperm are released into the embryo sac; one fertilises the ovule to produce a diploid zygote, and the other joins with two polar nuclei in the ovule to produce a triploid nucleus that will then develop into the nutrient-rich endosperm (Willmer, 2011). Pollen grain diameter is usually in the range 20-70 μm, and the surface structure and morphology varies considerably between plant species and dispersal mechanism (Wiltshire, 2010). Air temperature can have an effect on pollen formation and viability, with high temperatures potentially leading to sterile pollen (Bosland and Votava, 2012). Irradiated pollen grains are still able to germinate and produce pollen tubes that reach the ovule (Germana, 2012). Although they are unable to fertilise the egg cell, this process induces parthenogenesis and has been widely used to produce haploid fruits (Germana, 2012).

Slowing Development Facilitates Arabidopsis mgt Mutants to Accumulate Enough Magnesium for Pollen Formation and Fertility Restoration

Magnesium (Mg) is an abundant and important cation in cells. Plants rely on Mg transporters to take up Mg from the soil, and then Mg is transported to anthers and other organs. Here, we showed that MGT6+/− plants display reduced fertility, while mgt6 plants are fertile. MGT6 is expressed in the anther at the early stages. Pollen mitosis and intine formation are impaired in aborted pollen grains (PGs) of MGT6+/− plants, which is similar to the defective pollen observed in mgt5 and mgt9 mutants. These results suggest that Mg deficiency leads to pollen abortion in MGT6+/− plants. Our data showed that mgt6 organs including buds develop significantly slower and mgt6 stamens accumulate a higher level of Mg, compared with wild-type (WT) and MGT6+/− plants. These results indicate that slower bud development allows mgt6 to accumulate sufficient amounts of Mg in the pollen, explaining why mgt6 is fertile. Furthermore, we found that mgt6 can restore fertility of mgt5, which has been reported to be male sterile due to defects in Mg transport from the tapetum to microspores and that an additional Mg supply can restore its fertility. Interestingly, mgt5 fertility is recovered when grown under short photoperiod conditions, which is a well-known factor regulating plant fertility. Taken together, these results demonstrate that slow development is a general mechanism to restore mgts fertility, which allows other redundant magnesium transporter (MGT) members to transport sufficient Mg for pollen formation.

SPECTRUM OF CAUSATIVELY RELEVANT AEROALLERGENS OF SOUTHERN UKRAINE WHICH PROVOKE ALLERGIC REACTION IN PATIENTS WITH POLLINOSIS ON THE EXAMPLE OF KHERSON REGION

The aim: the analysis of spectrum of causatively relevant aeroallergens of southern Ukraine which provoke allergic reaction in patients with pollinosis. Materials and methods: ambulatory medical records data and skin prick-testing results of 477 patients afflicted with pollinosis in Kherson (250 women – 52,41 % and 227 men – 47,59 %) aged from 19 to 66 years old (38,05±0,45) have been analyzed retrospectively. Results: all patients revealed clinical implications mainly during the September-October period. Clinically 151 patients (31,66%) showed severe and 326 (68,34%) moderate to severe clinical progression. Composite family plants (composites) proved to be the most widespread plants (69,76 %) among all pollen aeroallergens of Kherson region. Weeds and sunflower showed the highest percentage among composites: Ambrosia topped to 79,87 %, Helianthus (sunflower)– 75,68 %, Artemisia – 42,98 %, and Cyclachaena (sump weed) – 36,48 %. Wild and cultivated grains’ aeroallergens toped to 28,22 %. Among them the sensibility to Lolium perenne (19,28 %), Festuca pratensis (17,82 %), Dactylis glomerata (17,82 %) has been noticed more often. Tree aeroallergens topped to 2,03 %. Conclusions: The third wave of pollen formation (September-October period) appeared to be the most significant and clinically hard for Kherson region. According to the specific allergologic research data, allergens of the composite family plants, among which pollen of Ambrosia, Helianthus, Artemisia and Cyclachaena cause more frequent sensibility, turned to be the most widespread causatively relevant aeroallergens in Kherson region which provoke allergic reaction in patients with pollinosis. The second place was taken by wild and cultivated grains’ aeroallergens, among which sensibility to Lolium perenne, Festuca pratensis, Dactylis glomerata proved to be the most common.

Morphology of flower and development of male reproductive sphere of plants Fagopyrum homotropicum Ohnishi

Вид гречихи Fagopyrum homotropicum Ohnishi стал известен научной общественности в 1995 году. Благодаря своей скрещиваемости с культурными видами гречихи он был включен в разнообразные селекционно-генетические программы исследований. При этом многие аспекты его репродуктивной биологии, в частности эмбриологические особенности, остаются на данный момент неизученными. Настоящая работа посвящена изучению морфологии цветка и процессов формирования мужской репродуктивной сферы растений F. homotropicum. В результате проведенных исследований показано, что цветок F. homotropicum представлен 9 различными вариантами строения по количеству органов. Типичный вариант строения P5A8G(3) встречался с частотой 93,4%. Варианты P5A6G(3), P5A7G(3), P5A8G(0), P6A8G(3) зафиксированы с частотой 1,0-2,2%. Цветки нетипичного строения F. homotropicum обычно имеют измененное количество органов андроцея или гинецея. Полиморфизм строения цветков, таким образом, характерен для всех видов cymosum группы. Процессы микроспорогенеза и микрогаметогенеза протекают у F. homotropicum аналогично тому, как они описаны для других изученных видов Fagopyrum. Количество материнских клеток микроспор в гнезде пыльника составило 5,2±0,15. Обнаружены также нарушения в ходе эмбриологических процессов: приостановка развития клеток спорогенной ткани с последующей ее дегенерацией. Контрастные погодные условия в период формирования пыльцы не оказали влияния на ее фертильность: в разные годы она составила 93,4 и 93,6%. The species of buckwheat Fagopyrum homotropicum Ohnishi became known to the scientific community in 1995. Due to its crossbreeding with the cultural species of buckwheat, it was included into several breeding and genetic research programs. At the same time, many aspects of its reproductive biology, in particular embryological features, remain unknown at the moment. The present work is aimed to the study of flower morphology and the processes of formation of the male reproductive sphere of the F. homotropicum plants. Here we show that the flowers F. homotropicum are represented by 9 different variants of the structure by the number of organs. A typical variant of the structure P5A8G(3) met with a frequency of 93,4 %. The variants P5A6G(3), P5A7G(3), P5A8G(0), P6A8G(3) are recorded with a frequency of 1,0-2,2 %. The flowers of the atypical structure of F. homotropicum usually have an altered number of androecium or gynaecium organs. Polymorphism of the structure of flowers, thus, is characteristic of all species of cymosum group. The processes of microsporogenesis and microgametogenesis take place in F. homotropicum in the same way as they were described for other studied Fagopyrum species. The number of microspore mother cells in the anther nest was 5,2 ± 0,15. Abnormalities were also found in embryological processes: the suspension of the development of cells of sporogenic tissue, followed by its degeneration. Contrast weather conditions during the pollen formation did not affect its fertility: in different years, it amounted to 93,4 and 93,6 %.

Millet pollen quality depending on the growing conditions and varietal characteristics

The aim of the study is to find out the features of forming the quality of pollen grains, namely their size and viability, depending on the growing conditions and varietal characteristics of switchgrass. The research deals with the peculiarities of millet pollen grains formation, depending on the conditions of vegetation and varietal composition, as the pollen quality – its size and viability – affects the seeds germination. It was established that the pollen quality depended both on the conditions of the growing season and varietal composition. Thus, in the 2018 growing season the weather conditions in the flowering phase (July) were favorable for the viable pollen formation, and high temperatures in August, exceeding 35 oC after 11:00 am at the absence of precipitation affected the viability of the pollen. In all the varieties, the viability of pollen was low. The most viable pollen was in the Morozko variety and the least viable – in the Cave-in-Rock variety, which negatively affected the seed germination. Pollen grains are spherical in form and colorless. They were not uniform in size both depending on the genotype and the growing conditions. In 2018 the pollen grains size ranged from 29 to 60 microns, in 2019 – from 5.2 to 57.2 microns. In 2019, the smaller pollen formed compared to 2018 due to high average daily air temperatures that exceeded average daily long-term values and due to significant moisture deficiency. Millet pollen is not homogeneous, depending on the both genotype and vegetation conditions. In 2018, which was favorable for pollen formation, the size varied from 29 to 60 microns, and in the less favorable 2019, it was smaller and ranged from 5.2 to 57.2 microns. The largest size of pollen grains formed in the Sunburst variety compared to other varieties. The most uniform – 85.3 % of the total amount – was the pollen in the Cave-in-Rock variety. Key words: pollen grains, quality, pollen size, viability, pollen form, variability, uniform.

ARABIDOPSIS POLLEN ABORTION MUTANT 2, Encoding an Unknown Protein with Three WD40-Repeats, Is Essential for Late Pollen Mature in Arabidopsis thaliana

Background In flowering plants, pollen formation is a very complex process. It is strictly regulated by various genetic factors. Some of these factors have been identified, but the regulatory mechanism concerning the process of late pollen maturing remains unknown.Results Previously we identified and reported an ARABIDOPSIS POLLEN ABORTION MUTANT 2 (APAM2) gene which was required for pollen formation in Arabidopsis thaliana. In this study, we further analysed the phenotype of atapam2 and the expression pattern of AtAPAM2. Mutation in AtAPAM2 drastically affected male gametophytic function, impacted the late pollen mature process, and resulted in mature pollen grains from atapam2/+ plants being shrivelled and dead. It was expressed among seedlings, roots, stems, leaves, siliques, flowers, and pollen grains. However, its expression levels in flowers and pollen grains were much higher than in other tissues. AtAPAM2 encodes an unknown protein with three WD40-repeats with localisation in plasma membrane and nucleus, and homologous proteins exist in many species.Conclusions We determined that AtAPAM2 may play a role in pollen formation and other development processes by interacting with other proteins, but we did not identify the protein that interacts with it.