MALE-STERILITY

Several forms of pollination control Manual emasculation Use of male sterility Use of self-incompatibility Use of male gametocides Use of genetically engineered “pollen killer” genetic system

Male-sterility Plant that do not produce viable, functional pollen grains An inability to produce or to release functional pollen as a result of failure of formation or development of functional stamens, microspores or gametes

Phenotypic classes of sterility “Pollen sterility” in which male sterile individuals differ from normal only in the absence or extreme scarcity of functional pollen grains (the most common and the only one that has played a major role in plant breeding) “Structural or staminal male sterility” in which male flowers or stamen are malformed and non functional or completely absent “Functional male sterility” in which perfectly good and viable pollen is trapped in indehiscent anther and thus prevented from functioning

Types of Male-sterility Genetic male sterility Cytoplasmic male sterility Environment sensitive genetic male sterility Chemical induced male sterility Genetically engineered male sterility

Cytoplasmic male-sterility CMS is the result of mutation in the mitochondrial genome (mtDNA), which leads to mitochondrial dysfunction. Stamen (anther and filament) and pollen grains are affected

Cytoplasmic male-sterility The nuclear genetic control of CMS is predominantly governed by one or more recessive genes, but can be also dominant genes as well as polygenes The different mtDNA restriction endonuclease digestion patterns are reflections of aberrant intra- or inter molecular DNA recombination events in the mitochondrial genome which have either modified existing genes or related new genes some of which are more or less related to the male sterile phenotypes

Cytoplasmic male-sterility Origins: 1. Intergeneric crosses 2. Interspecific crosses 3. Intraspecific crosses 4. Mutagens (EMS, EtBr) 5. antibiotic (streptomycin and Mitomycin) 6. Spontaneus

CMS Characterization It has been traditionally characterized by the restore genes required to overcome the CMS and to provide male sterile progeny in the male sterile system CMS restoration is by nuclear genes, frequently dominant in action, in many cases, few in number The CMS restore genes temporarily suppress the expression of the CMS permitting normal or near-normal pollen production

Types a. Autoplasmic CMS has arisen within a species as a result of spontaneous mutational changes in the cytoplasm, most likely in the mitochondrial genome b. Alloplasmic CMS has arisen from intergeneric, interpecific or occasionally intraspecific crosses and where the male sterility can be interpreted as being due to incompatibility or poor co-operation between nuclear genome of one species and the organellar genome another CMS can be a result of interspecific protoplast fusion

CMS mechanism of action Abnormal behavior of the tapetum in the anther Genetic determinant of CMS reside in mitochondria Nuclear gene control the expression of CMS

CMS Limitations Pleiotropic negative effect of the CMS on agronomic quality performance of plants in the CMS cytoplasm Enhanced disease susceptibility Complex and environmentally unstable maintenance of male sterility and/or male fertility restoration Inability to produce commercial quantities of hybrid seed economically because of poor floral characteristic of cross pollination

CMS Utilization It provides a possible mechanism of pollination control in plants to permit the easy production of commercial quantities of hybrid seeds It consists of a male sterile line (the A-line), an isogenic maintainer line (The B line), and if necessary also restore line (the R-line) A lines are developed by back-crossing selected B-lines to a CMS A-line for 4 – 6 times to generate a new A-line, B and R-lines are developed by similar back cross procedures using a CMS R-line as female in the original cross and a new line as the recurrent parent in 4 – 6 backcrosses

CMS Utilization Selfing the last backcross generation two successive times and selection of pure breeding male fertility restore line is required to complete the development of the new R-lines developed in the CMS Current commercial hybrid seed production relies entirely on the block method (alternating strips of female and male genotypes

Fertility restoration in maize

Simple hybrid with cms and restoration rr S rr F CMS line (A-line) CMS, rfrf x Maintainer line (B-line) N, rfrf Large amounts of CMS line rr S F x RR Male line (C-line) N and RfRf Rr S Fertile F1 hybrid CMS, Rfrf

Nuclear male sterility Originated through spontaneous mutation or mutation by ionizing radiation and chemical mutagens such as ethyl methane sulphonate (EMS) and ethyl imine (EI) or by genetic engineering, protoplast fusion can probably be found in all diploid species Usually controlled by mutations in genes in the single recessive genes affect stamen and pollen development, but it can be regulated also by dominant genes

Morphology Variable (complete absence of male reproductive organs to the formation of normal stamen with viable pollen that fail to dehisce) It is not distinguishable from parent fertile plants with the exception of flower structure Male sterile flowers are commonly smaller in size in comparison to the fertile The size of stamens is generally reduced

Determining factor Temperature (TGMS) Photoperiod (PGMS) Changing the optimal temperature can induce sterility (23°C) Photoperiod (PGMS) It has a strong influence (Photoperiod sensitive) Changing the growth habit can stimulate the sterility (23°C - 29°C)

Cytological Changes Breakdown in microsporogenesis can occur at a number of pre-or postmeiotic stages The abnormalities can involve aberration during the process of meiosis, in the formation of tetrads, during the release of tetrad (the dissolution of callose), at the vacuolate microspore stage or at mature or near-mature pollen stage

Use of genic male sterility in hybrid programs Male sterile plants of monoecious or hermaprodite crops are potentially useful in hybrid program because they eliminate the labor intensive process of flower emasculation

Hybrid seed production with GMS and restoration Male sterile line X Male fertile line msms MsMs Male sterile line X Maintainer line msms Msms Seed for harvested in bulk from male sterile line

Maintenance plot Hybrid seed production plot Plants with Msms and msms genotypes Female rows male rows Female rows Msms & msms Msms & msms Harvest seed only from sterile plants Remove fertile plants from rows before anthesis, harvest seed from sterile plants

Cytoplasmic-genetic male sterility A case of cytoplasmic male sterility where a nuclear gene for restoring fertility in the male sterile line is known. The fertility restorer gene R is dominant.

Various Genotypes and Phenotypes rr S Cytoplasm sterile Nuclear gene non restorer rr F Cytoplasm fertile Nuclear gene non restorer RR S Cytoplasm sterile (Male fertile) Nuclear gene restorer in homozygous RR or heterozygous Rr state The effect of sterile cytoplasm is negated by the restorer gene Rr S

CHEMICAL INDUCED MALE-STERILE

Biochemical means of producing male sterile plants Feminizing hormones Inhibitors of anther or pollen development Inhibitors of pollen fertility

Chemical hybridizing agent (CHA) Could be used in the large scale commercial production of hybrid seed Are applied to plant only at certain critical stage of male gametophyte development

The logic of chemical hybridization High degree of efficacy and developmental selectivity Persistence during the development of flower or spikes Low cost Acceptable levels of toxicity to people and the environment Low general phytotoxicity Agronomic performance of hybrid seed produced is not inferior to equivalent crosses produced by genetic methods

CHAs and pollen development There are at least 4 classes of chemical agents: a. Plant growth regulators and substances that disrupt floral development b. Metabolic inhibitors c. inhibitors of microspore development d. inhibitors of pollen fertility

Plant growth regulators and substances that disrupt floral development Plant hormones/hormones antagonists a. auxins and auxin antagonists (NAA, IBA, 2,4-D, TIBA, MH) b. Gibberellins and antagonist (GA3, GA4+7, CCC: 2-chloroethyl-trimethyl ammonium chloride) c. Abscisic acid Other substances a. LY195259 b. TD1123

Metabolic Inhibitors There are halogenated aliphatic acids (alpha, beta-dichloroisobutyrate and 2,2-dichloropropionate salts) and arsenicals (methanearsonate salts) They affect mitochondrial protein by reducing the efficiency of normal metabolic processes

Inhibitors of microspore development Copper chelators Ethylene Fenridazon Phenylcinnoline carboxylates (SC-1058, SC-1271 and SC-2053)

Inhibitors of pollen fertility Azetidine-3-carboxylate (A3C, CHA™)