Construction of backcrossedGelidium male-sterile and male-fertile lines and their growth comparison

The male sterility gene from a male-sterile, green,Gelidium vagum line was introduced to a wild-type line through repeated backcrossing and selection for five generations. The plants from the recurrent parent, the male-sterile green, the backcross-5 fertile and the backcross-5 male-sterile lines were compared for their growth performance. The backcross-5, red, male-sterile plants grew at a significantly higher rate than the plants from other lines, suggesting that male-sterile gametophytes would likely be better candidates for aquaculture than normal wild-type plants of this species. This study also provides evidence that the growth rate ofG. vagum decreases as a consequence of reproduction.     

MS1 is essential for male fertility by regulating the microsporocyte cell plate expansion in soybean

Male sterility is an essential trait in hybrid seed production, especially for monoclinous and autogamous food crops. Soybean male-sterile ms1 mutant has been known for more than 50 years and could be instrumental in making hybrid seeds. However, the gene responsible for the male-sterile phenotype has remained unknown. Here, we report the map-based cloning and characterization of the MS1 gene in soybean. MS1 encodes a kinesin protein and localizes to the nucleus, where it is required for the male meiotic cytokinesis after telophase Ⅱ. We further substantiated that MS1 colocalizes with microtubules and is essential for cell plate formation in soybean male gametogenesis through immunostaining. Both ms1 and CRISPR/Cas9 knockout mutants show complete male sterility but are otherwise phenotypically normal, making them perfect tools for producing hybrid seeds.The identification of MS1 has the practical potential for assembling the sterility system and speeding up hybrid soybean breeding.     

Strong sexual selection in males against a mutation load that reduces offspring production in seed beetles

Theory predicts that sexual reproduction can increase population viability relative to asexual reproduction by allowing sexual selection in males to remove deleterious mutations from the population without large demographic costs. This requires that selection acts more strongly in males than females and that mutations affecting male reproductive success have pleiotropic effects on population productivity, but empirical support for these assumptions is mixed. We used the seed beetle Callosobruchus maculatus to implement a three‐generation breeding design where we induced mutations via ionizing radiation (IR) in the F₀ generation and measured mutational effects (relative to nonirradiated controls) on an estimate of population productivity in the F₁ and effects on sex‐specific competitive lifetime reproductive success (LRS) in the F₂. Regardless of whether mutations were induced via F₀ males or females, they had strong negative effects on male LRS, but a nonsignificant influence on female LRS, suggesting that selection is more efficient in removing deleterious alleles in males. Moreover, mutations had seemingly shared effects on population productivity and competitive LRS in both sexes. Thus, our results lend support to the hypothesis that strong sexual selection on males can act to remove the mutation load on population viability, thereby offering a benefit to sexual reproduction.     

Genetics and cytology of a new genic male-sterile soybean [Glycine max (L.) Merr.]

 Genetic and cytological studies were conducted with a new male-sterile, female-fertile soybean [Glycine max (L.) Merr.] mutant. This mutant was completely male sterile and was inherited as a single-recessive gene. No differences in female or male gamete transmission of the recessive allele were observed between reciprocal cross-pollinations in the F₁ or F₂ generations. This mutant was not allelic to any previously identified soybean genic male-sterile mutants: ms1, ms2, ms3, ms4, ms5, or ms6. No linkage was detected between sterility and flower color (W1 locus), or between sterility and pubescence color (T1 locus). Light microscopic and cytological observations of microsporogenesis in fertile and sterile anthers were conducted. The structure of microspore mother cells (MMC) in male-sterile plants was identical to the MMCs in male-fertile plants. Enzyme extraction analyses showed that there was no callase activity in male-sterile anthers, and this suggests that sterility was caused by retention of the callose walls, which normally are degraded around tetrads at the late tetrad stage. The tapetum from male-sterile anthers also showed abnormalities at the tetrad stage and later stages, which were expressed by an unusual formation of vacuoles, and by accumulation of densely staining material. At maturity, anthers from sterile plants were devoid of pollen grains. Received: 13 May 1996 / Revision accepted: 19 August 1996     

A straight-forward seed production technology system for foxtail millet (Setaria italica)

For autogamous crops, a precondition for using heterosis is to produce sufficient pure male-sterile female parents that can be used to produce hybrid seeds. To date, cytoplasmic male sterility(CMS)and environment-sensitive genic male sterility(EGMS) have been used commercially to exploit heterosis for autogamous species. However, neither CMS nor EGMS has been established for foxtail millet(Setaria italica). Here, we report on the establishment and application of a seed production technology(SPT)...     

The impacts of Wolbachia and the microbiome on mate choice in Drosophila melanogaster

Symbionts and parasites can manipulate their hosts’ reproduction to their own benefit, profoundly influencing patterns of mate choice and evolution of the host population. Wolbachia is one of the most widespread symbionts among arthropods, and one that alters its hosts’ reproduction in diverse and dramatic ways. While we are beginning to appreciate how Wolbachia's extreme manipulations of host reproduction can influence species diversification and reproductive isolation, we understand little about how symbionts and Wolbachia, in particular, may affect intrapopulation processes of mate choice. We hypothesized that the maternally transmitted Wolbachia would increase the attractiveness of its female hosts to further its own spread. We therefore tested the effects of Wolbachia removal and microbiome disruption on female attractiveness and male mate choice among ten isofemale lines of Drosophila melanogaster. We found variable effects of general microbiome disruption on female attractiveness, with indications that bacteria interact with hosts in a line‐specific manner to affect female attractiveness. However, we found no evidence that Wolbachia influence female attractiveness or male mate choice among these lines. Although the endosymbiont Wolbachia can greatly alter the reproduction of their hosts in many species, there is no indication that they alter mate choice behaviours in D. melanogaster.     

Gene expression and the control of spermatid morphogenesis in Drosophila melanogaster.

The genetic control of spermatid morphogenesis was studied by light microscopy through the analysis of meiotic and premeiotic lesions. Sperm disfunction-type male-sterile mutations were screened for novel “early effect” mutations: (1) timing mutations, in which mitochondrial aggregation occurs before instead of after meiosis; (2) mutations which affect the spindle structure, e.g., a mutant with second-division monoastral spindle; (3) mutations which cause deformations in primary spermatocyte structures. It is shown, in addition to the examples cited above, that normal meiosis may often serve as an early marker for normal differentiation, and that approximately 20% of male-sterile mutations are meiotic mutants. The role of the Y chromosome was reexamined. The interaction between Y factors and X-linked male steriles is in many cases additive, indicating that Y gene products are essential for normal development of the primary spermatocytes. Furthermore, XO males are shown to be extreme meiotic mutants. It is argued that spermatid morphogenesis is totally dependent on developmental processes in the primary spermatocyte stage. The relations among developmental processes in early spermatogenesis are discussed in terms of gene activity.     

Anent the Genomics of Spermatogenesis in Drosophila melanogaster

An appreciable fraction of the Drosophila melanogaster genome is dedicated to male fertility. One approach to characterizing this subset of the genome is through the study of male-sterile mutations. We studied the relation between vital and male-fertility genes in three large autosomal regions that were saturated for lethal and male-sterile mutations. The majority of male-sterile mutations affect genes that are exclusively expressed in males. These genes are required only for male fertility, and several mutant alleles of each such gene were encountered. A few male-sterile mutations were alleles of vital genes that are expressed in both males and females. About one-fifth of the genes in Drosophila melanogaster show male-specific expression in adults. Although some earlier studies found a paucity of genes on the X chromosome showing male-biased expression, we did not find any significant differences between the X chromosome and the autosomes either in the relative frequencies of mutations to male sterility or in the frequencies of genes with male-specific expression in adults. Our results suggest that as much as 25% of the Drosophila genome may be dedicated to male fertility.     

The role of X-linked lethal and viable male-sterile mutations in male gametogenesis of Drosophila melanogaster: genetic analysis.

Summary The possibility that viable male-sterile mutations occur in vital genes and the role played by lethal mutations and viable male-steriles in male gametogenesis were studied. Five sterile loci were identified among the 30 most proximal vital loci of the X-chromosome and two of them were shown to be allelic with lethal mutations. Fertility tests on gynanders for nonautonomous lethal mutations proved that vital genes operate autonomously in male gonads, independently of their effect on somatic tissues. Fertility tests of ts lethals, shifted to the nonpermissive temperature after the TSP, showed that 40% of vital genes function in male gonads. It is further shown that about the same proportion of vital genes is operating in female gonads and that the two groups overlap by about 70%. The role of viable and lethal male sterile mutations in the control and regulation of male gametogenesis is discussed in detail.This work was supported by the Israel Commission for Basic Sciences.     

Characterization of three male-sterile mutants of Arabidopsis thaliana exhibiting alterations in meiosis

Male-sterile mutants are being studied to deepen our understanding of the complex processes of microsporogenesis and microgametogenesis. Due to difficulties associated with isolating the mutated gene, there is currently very little molecular information on the defects responsible for male sterility. As a first step in utilizing male-sterile mutants to better understand the bio-chemical and molecular processes that control pollen development, we have characterized a number of Arabidopsis thaliana lines that were generated by seed transformation and exhibit male sterility. We report here the identification and characterization of three male-sterile A. thaliana lines, all of which are tagged with T-DNA and show aberrant meiosis. A detailed cytochemical study was conducted on these lines to better understand the timing and nature of each mutation and to investigate how these mutations affect subsequent steps of pollen development. All three mutants undergo apparently normal morphogenesis until the onset of meiosis. In one line (6492) the mutation is most notable at the tetrad stage when up to eight microspores can be seen in each callose-encased tetrad. The resulting mutant microspores are of variable sizes and contain different amounts of DNA. Two other mutants (7219 and 7593) possess many common features, including variable developmental pathways, failure to produce callose, production of vacuolate, coenocytic (multi-nucleate) cells that are surrounded by persistent microsporocyte walls, and asynchronous patterns of development. Unlike the situation in wild-type plants, where developmental stages are correlated with bud length, such correlations are almost impossible with these two mutants. The sporogenous tissue within all three of these mutant lines collapses prior to anthesis.     

Isolation and genetic characterization of self-sterile mutants in a monoecious red alga Gelidium vagum (Gelidiales,Rhodophyta)

Sporelings of the monoecious red alga Gelidium vagum were placed into 4500 individual cultures after treatment with the chemical mutagen nitrosoguanidine and raised to sexual maturity to search for reproductive mutants. Isolates undergoing normal self-fertilization were discarded, leaving approximately 250 self-sterile plants and mutants with abnormal reproduction or reproductive structures. Self-sterile mutants were tested further in crosses to a fertile green marker stock. From the results, many mutants appeared to be either male-sterile, female-sterile or unable to form carposporophytes. Although sufficient data on the inheritance pattern of the self-sterile mutants are available for only a few of the isolates, some apparently stable sterility mutations with simple Mendelian transmission were identified. Preliminary testing of one of the male-sterile mutations confirmed that it effectively eliminated self-fertilization and facilitated the formation of hybrid plants in crosses.     

The combined effects of pre‐ and post‐copulatory processes are masking sexual conflict over mating rate in Gerris buenoi

In polygynandrous animals, post‐copulatory processes likely interfere with precopulatory sexual selection. In water striders, sexual conflict over mating rate and post‐copulatory processes are well documented, but their combined effect on reproductive success has seldom been investigated. We combine genetic parentage analyses and behavioural observations conducted in a competitive reproductive environment to investigate how pre‐ and post‐copulatory processes influence reproductive success in Gerris buenoi Kirkaldy. Precopulatory struggles had antagonistic effects on male and female reproductive success: efficiently gaining copulations was beneficial for males, whereas efficiently avoiding copulations was profitable for females. Also, high mating rates and an intermediate optimal resistance level of females supported the hypothesis of convenience polyandry. Contrary to formal predictions, high mating rates (i.e. the number of copulations) did not increase reproductive success in males or decrease reproductive success in females. Instead, the reproductive success of both sexes was higher when offspring were produced with several partners and when there were few unnecessary matings. Thus, male and female G. buenoi displayed different interests in reproduction, but post‐copulatory processes were masking the effects of copulatory mating success on reproductive success. Given the high mating rates observed, sperm competition could easily counter the effect of mating rates, perhaps in interaction with cryptic female choice and/or fecundity selection. Our study presents a complex but realistic overview of sexual selection forces at work in a model organism for the study of sexual conflict, confirming that insights are gained from investigating all episodes in the reproduction cycle of polygynandrous animals.     

GmMs1 encodes a kinesin-like protein essential for male fertility in soybean (Glycine max L.)

The application of heterosis is a promising approach for greatly increasing yield in soybean (Glycine max L.). Nuclear male sterility is essential for hybrid seed production and the utilization of heterosis. Here we report the cloning of the gene underlying the soybean male-sterile mutant ms-1, which has been widely used for recurrent selection in soybean breeding programs. We initially delimited the ms1 locus to a 16.15 kb region on chromosome 13, based on SLAF_BSA sequencing followed by genotyping of an F₂ population segregating for the locus. Compared with the same region in fertile plants, the mutant region lacks a sequence of approximately 38.7 kb containing five protein-coding genes, including an ortholog of the kinesin-like protein gene NACK2, named GmMs1. The GmMs1 knockout plants generated via CRISPR/Cas-mediated gene editing displayed a complete male-sterile phenotype. Metabolic profiling showed that fertile anthers accumulated starch and sucrose normally, whereas sterile anthers had higher anthocyanin levels and lower flavonoid levels and lower antioxidant enzyme activities. These results provide insights into the molecular mechanisms governing male sterility and demonstrate that GmMs1 could be used to create male-sterile lines through targeted mutagenesis. These findings pave the way for designing seed production technology and an intelligent male-sterile line system to utilize heterosis in soybean.     

Mutations leading to nuclear restoration of fertility in S male-sterile cytoplasm in maize

Summary Among the offspring of crosses involving S male-sterile shrunken-2 inbred lines and their corresponding isogenic maintainer lines a number of exceptional male-fertile plants were identified. Some of these were plants with entirely fertile tassels but most were chimeras involving both sterile and fertile tassel elements. The majority of male-fertile exceptional plants, upon crossing with male-sterile testers, produced male-sterile test-cross progeny, indicating that the male-fertile trait is not pollen transmissible. However, there were four separate instances, involving three of the inbred lines, in which the crosses with S male-sterile testers produced male-fertile progeny, indicating that the newly arisen male-fertile trait is pollen transmissible. In three of these cases, the male fertility can be traced to a single plant in essentially male-sterile families. The fourth evidently involved a change in a maintainer plant whose progeny thereafter segregated for the ability to restore S sterile cytoplasm. In all cases, the results of progeny tests are consistent with the gametophytic pattern of restoration associated with S male-sterile cytoplasm.The male-fertile exceptions described here can be accounted for formally as mutations at one or more restorer gene loci in the nucleus. Taking account of the fact that mutations of restorer genes have not been reported previously in maize, and that four such changes were encountered in the same strains in which we have identified other male-fertile exceptions involving change in the cytoplasm, we have suggested a common basis for the two kinds of events. According to this scheme, given the first appearance, by whatever process, of the male-fertile element in sterile cytoplasm, it may become established and continue to propagate either in the cytoplasm or in the nucleus. In the former case, the change registers as cytoplasmic and the new strain has the characteristics of a maintainer which transmits the male-fertile trait through the egg, but not the sperm; in the latter case, the change occurs in the nucleus and the new strain, now behaving as a restorer, transmits male fertility through both egg and sperm.Dedicated to Dr. M. M. Rhoades on the occasion of his 70th birthday.Research supported in part by CSRS grant 177-15-04, PL 89–106, and by NSF GB 29488.     

<Emphasis Type="Italic">Multiple impairments in male reproduction 1 (mimr1)</Emphasis>, a novel male-sterile mutant of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>, shows several defects in male reproductive development

We have characterized a new male-sterile mutant in Arabidopsis that exhibits conditional sterility but has restored fertility when drought-stressed. This mutant, multiple impairments in male reproduction 1 (mimr1), shows pleiotropic defects in both vegetative and reproductive development. Examination with dissecting and scanning electron microscopes revealed that its pollen grains are not effectively released from the anther locule after dehiscence, and anther differentiation is defective. Growth of the style and stamen filaments are also abnormal. Histological analysis demonstrated that these phenomena are due not only to a noticeably reduced extension of the stamen but also greater elongation of the pistil. Genetic analysis indicated that mimr1 is a single locus recessive nuclear mutant. The mutation can be mapped to a locus strongly linked to a 1200-kb region on Chromosome 3. Meta-analysis of expression patterning presented several candidate genes in that region. No mutants with similar phenotypes have previously been reported, suggesting that mimr1 is a novel male-sterile locus. Characterization of MIMR1 will provide further insights into the molecular basis for the development of plant reproductive organs.     

Molecular mapping of 36 soybean male-sterile, female-sterile mutants

Mutability of the w ( 4 ) flower color locus in soybean [Glycine max (L.) Merr.] is conditioned by an unstable allele designated w ( 4 ) -m. Germinal revertants, purple-flower plants, recovered among self-pollinated progeny of mutable flower plants were associated with the generation of necrotic root, chlorophyll-deficiency, and sterility mutations. Thirty-seven male-sterile, female-sterile mutant lines were generated from 37 independent reversion events at the w ( 4 ) -m locus. The first germinal revertant study had one male-sterile, female-sterile mutant (st8, T352), located on Molecular Linkage Group (MLG) J. The second study had 36 germinal-revertant derived sterility mutants descended from four mutable categories of w ( 4 ) -m. The mutable categories were designated; (1) low frequency of early excisions, (2) low frequency of late excisions, (3) high frequency of early excisions, and (4) high frequency of late excisions. The objectives of the present study were to; (1) molecularly map the 36 male-sterile, female-sterile mutants, and to (2) compare map locations of these mutants with T352 (st8), identified from the first germinal revertant study. Thirty-three of 36 male-sterile, female-sterile mutations were derived from germinal reversions that were classified in the late excision categories. Thirty-five male-sterile mutants mapped to the st8 region on MLG J. The only exception mapped to MLG G. Most likely mutants were generated through insertion of a putative transposon that was excised from the w ( 4 ) locus. The location of 36 of 37 mutations to a single chromosomal region suggests preference for sequence-dependent insertion.     

Mitochondrial Genome Dynamics in Plants and Animals: Convergent Gene Fusions of a MutS Homologue

Mitochondrial processes influence a broad spectrum of physiological and developmental events in higher eukaryotes, and their aberrant function can lead to several familiar disease phenotypes in mammals. In plants, mitochondrial genes directly influence pollen development and the occurrence of male sterility in natural plant populations. Likewise, in animal systems evidence accumulates to suggest important mitochondrial functions in spermatogenesis and reproduction. Here we present evidence for a convergent gene fusion involving a MutS-homologous gene functioning within the mitochondrion and designated Msh1. In only plants and soft corals, the MutS homologue has fused with a homing endonuclease sequence at the carboxy terminus of the protein. However, the endonuclease domains in the plants and the soft corals are members of different groups. In plants, Msh1 can influence mitochondrial genome organization and male sterility expression. Based on parallels in Msh1 gene structure shared by plants and corals, and their similarities in reproductive behavior, we postulate that this convergent gene fusion might have occurred in response to coincident adaptive pressures on reproduction. [Reviewing Editor: Dr. Deborah Charlesworth]     

Downy mildew incidence of pearl millet hybrids with different male-sterility inducing cytoplasms

The use of different sources of cytoplasmic male sterility (CMS) in hybrid seed production of pearl millet [Pennisetum glaucum (L.) R. Br.] is advocated to avoid possible disease epidemics occurring due to cytoplasmic uniformity. The effects of commercially unexploited, but potentially exploitable, sources of CMS, like A₂, A₃ and A₄, on downy mildew [Sclerospora graminicola (Sacc.) Schroet] incidence were studied by using the disease incidence of isonuclear hybrids with male-sterile and fertile cytoplasm. The mean downy mildew incidence of hybrids carrying different male-sterile cytoplasm was similar to that of hybrids retaining the fertile cytoplasm. The cytoplasm accounted for only 0.6% of the total variation and its effect was non-significant; pollinators could explain most of the variation in determining the disease incidence of hybrids. This suggested that these male-sterile cytoplasms are not linked to downy mildew susceptibility and thus can be exploited commercially to broaden the cytoplasmic base of the male-sterile lines and, ultimately, of hybrids.     

Cytoplasmic male sterility in the olive (Olea europaea L.)

The olive tree is usually hermaphrodite but self-incompatible. In the Western Mediterranean some cultivars are totally male-sterile. Three different male-sterile phenotypes have been recognised. To infer the genetic basis of male sterility we studied its inheritance and cytoplasmic diversity in wild (oleaster) and cultivated Mediterranean olive. In the cross Olivière×Arbequina, the male-sterile trait was maternally inherited and affected all progenies. We also checked that both chloroplast and mitochondrial DNAs are maternally inherited. RFLP studies on chloroplast and mitochondrial DNAs revealed several cytotypes: two chlorotypes and four mitotypes in cultivars and oleaster (wild or feral Mediterranean olive). Furthermore, a total linkage desequilibrium between the CCK chlorotype and the MCK mitotype in cultivars and oleaster from different regions supports the fact that paternal leakage of organelles was not observed. The male sterility (ms 2) displayed by Olivière, plus six other cultivars and three oleaster was strictly associated with the CCK chlorotype and the MCK mitotype. These facts suggest that Olivière carries cytoplasmic male sterility. Male-fertile and male-sterile oleasters carrying this cytotype showed the presence of restorer alleles. This CMS might be due to a distant cross between olive taxa. The two other male-sterile phenotypes displayed by Lucques (ms 1) and Tanche (ms 3) were associated with the ME1 mitotype but we have not demonstrated CMS. Received: 26 July 1999 / Accepted: 27 August 1999     

A new and distinctive male-sterile,female-fertile desynaptic mutant in soybean (Glycine max)

A spontaneous desynaptic mutation, affecting only microsporogenesis and causing pollen sterility, has been detected in BR97-12986H, a line of the official Brazilian soybean breeding program. In this male-sterile, female-fertile mutant, up to metaphase II, the meiotic behavior was similar to that described for the st series of synaptic mutants previously reported in soybean. Besides many univalents, few or total absence of bivalents were recorded in diakinesis. Bivalents presented one or two terminal chiasmata, while univalents retained the sister chromatid cohesion. Bivalents and most univalents congregated at the equatorial metaphase plate, although univalents frequently migrated to the poles prematurely. Laggards resulting from delay in chiasmata terminalization were also recorded. Distinctly different in their behavior from st series soybean mutants, telophase I-originated micronuclei of different sizes organized their own spindle in the second division. This behavior contributed towards an increase in genome fractionation. Several microspores and microcytes of different sizes were recorded at the end of meiosis. Pollen sterility was estimated at 91.2%. Segregation ratio for sterility in this line and its progenies reached 3:1. Allelism tests with st series of synaptic mutants are in progress. The importance of male-sterile, female-fertile mutations for soybean breeding programs is discussed.