Mechanism of nuclear male sterility gene and its expression

1. GPB 605 Advanced Plant Breeding Systems
Nuclear Male Sterility
Sri Subalakhshmi V K I
2019608015
I Ph.D (GPB)

2. Introduction
• Nuclear male sterility is also known as genetic male sterility.
• Governed by single recessive gene ms
• Dominant gene governing in safflower
• ms alleles may affect staminal initiation, stamen or anther sac
development, PMC formation, meiosis, pollen formation,
maturation and dehiscence.

3. Types
 Environment insensitive GMS: ms gene expression is much
less affected by the environment.
 Environment sensitive GMS: ms gene expression occurs
within a specified range of temperature and /or photoperiod
regimes (Rice, Tomato, Wheat etc.).

4. TGMS
 sterility is at particular temperature e.g. In rice TGMS line
(Pei- Ai645) at 23.30C (China).
 TGMS at high temperature is due to failure of pairing of two
chromosomes was evident.
 This abnormality leads to abnormal meiosis, abnormal or sterile
pollens.
 Anthers were shriveled and non-dehiscence-Male sterile.
 However, these lines produced normal fertile pollen at low temp.
 Sensitive period : PMC formation to Meiosis

5. 2. PGMS: Governed by 2 recessive
genes.
 Sterility is obtained in long day
conditions while in short days,
normal fertile plant.
 Rice:- Sterile under Long day
conditions (13 hr. 45 min +
Temp. 23-290 C) but fertile
under short day conditions.
 Sensitive period: Differentiation
of secondary rachis branches to
PMC formation

6. Nuclear male sterility hybrid seed
production

7. Case study

8. • ms mutant represents a new locus for NMS, as male sterile 9
(ms9) and the casual gene as Ms9.
• male sterility of ms9 is stable under variation in day length and
temperature conditions, and that the mutation is fully
penetrant.

9. • Wild BTx623 – protrusion of anthers
from spikelets and stigma is partially
visible as they are surrounded by fluffy
anthers.
• ms 9 – white hairy stigma is visible as
no anthers are protruded out. It has
only small pale anthers
• Female parts of both wild and mutant
remains same
• Difference observed was no viable
pollen formation in ms 9 mutant.

10. • Normal seed set was observed in manually pollinated BC1F2
ms 9 panicles
• Lack of seeds observed in bagged panicles as it lacks fertile
pollen source and unable to produce seeds.

11. • Wild BTx623 anther lobes were
full of round, reddish, mature
pollen grains
• the anther lobes of ms9 mutants
were empty
• A microscopic view of ms9 anther
showed no sign of mature pollen
being released

12. • WT pollen became round, and the tapetum and endothecium layers
degenerated completely
• microspores in ms9 anther lobes degenerated completely, resulting in
little or no cell debris within the empty anthers at pollen maturation
stage
During pollen
maturation stages

13. Findings…
• 1:1 segregation pattern observed and this concluded that ms9 is
recessive mutation in single nuclear gene.
• deleterious mutation located in Sobic.002G221000 gene encoding a
PHD-finger transcription factor.
• C-to-T transition at nucleotide 1207 in Sobic.002G221000 gene in
ms9 causes an amino acid change from arginine to tryptophan at
conserved position 218
• Sobic.002G221000 is Ms9 in sorghum.
• Ms9 gene has similar expression pattern as Ms1 in Arabidopsis and
Ptc1 in rice, 100% identical to those of Ms7 in maize

14. • The Ms3 allele for DMS was isolated through ethyl
methanesulfonate (EMS)-induced mutagenesis from ‘Chris’
hard red spring wheat in Aegilops tauschii

15. Findings…
• stable within the normal range of greenhouse temperatures
for wheat (16–25 ◦C)
• Ms3 was localized close to the centromere on the short arm
of chromosome 5A
• association analysis identified two SNP tags with nearly
perfect association with male sterility, S5A_138430506 and
S5A_266997650.

16. • the low recombination rates near the centromere can be an
advantage for marker-assisted selection.
• In contrast with recessive male sterility systems, it is not
necessary to complete a generation of selfing to obtain true-
breeding fertile genotypes.
• Because progeny of DMS (Ms3/ms3) plants pollinated by a
fertile (ms3/ms3) male parent segregate 1:1 for the presence
of the Ms3 sterility allele, sterile individuals are readily
retained in the population.

17. su
• Nongken 58S (NK58S) is the first spontaneous PGMS mutant
found in 1973 from the japonica (O. sativa ssp. japonica)
cultivar Nongken 58 (NK58)

18. • NK58S retains complete male sterility - day length
(photoperiod) is longer than 13.75 h during the anther
development at temp ~29 °C
• Peiai 64S (PA64S) is NK58S-derived line with indica (O. sativa
ssp. indica) genetic background and exhibits male sterility at
temperatures >23.5 °C during the anther development
• F2 population showed 3:1 segregating ratio (880 F:105 S) -
TGMS in PA64S is determined by one major recessive locus

19. • p/tms12-1 confers PGMS in NK58S and TGMS in PA64S .
• Locus encodes a noncoding RNA precursor that produces a
21-nt small RNA.
• Transformation of NK58S and PA64S plants with the allele
P/TMS12-1 produced the wild-type small RNA (osa-
smR5864w) and suppressed both PGMS and TGMS under
long-day and high-temperature conditions.
• 5.8-kb region of the locus is mapped on chromosome 12.
• As the locus pms3 conferring PGMS in NK58S has been
mapped to the region of chromosome 12

20. • Sequence analysis revealed a C-to-G mutation in region of
PA64S and its donor japonica parent line NK58S.
• the small RNA osa-smR5864w encoded by P/TMS12-1 leads to
suppression of the PGMS and TGMS traits
• Only one SNP of the marker PASNP3 was identified, with a C
residue in all the fertile lines and a G residue in all the PGMS
and TGMS lines
• the point mutation in p/tms12-1 may lead to a loss of function
of osa-smR5864m and confer PGMS and TGMS in the japonica
and indica genetic backgrounds

21. • differences in gene expression between the ‘Dong A’ male
sterile mutant and its wild type (WT) was studied
• gene expression patterns were compared at three
developmental stages of the WT and GMS mutant

22. • At one day post anthesis
• WT – normal floral
phenotypes with deeply
stained pollen grains
• Mutant – abnormal floral
phenotypes with shorter
filaments and no pollen grains
were stained
• Little nutritional materials
were accumulated

23. • WT – tapetal cells were
stained strongly and
ubisch bodies extruded
(meiosis);
• tapetal cells slightly
vacuolated and exine
formation (tetrad);
• extine thicker & deeply
stained (uninucleate
microspores)
• sporopollenin deposition
occurs normally
• Mutant – tapetal layer is less stained
and highlt vacuolated with abnormal
degradation.
• No ubisch bodies,
• thin extine layer
• Sporopollenin synthesis was deficient

24. • 21,503 to 37,352 genes were detected in WT and GMS mutant
anthers.
• Compared with WT, 9,595 (30% of the expressed genes),
10,407 (25%), and 3,139 (10%) genes were differentially
expressed at the meiosis, tetrad, and uninucleate microspore
stages of GMS mutant anthers, respectively.

25. • genes involved in histone modification and DNA methylation,
hormone signaling, carbon and energy metabolism, pollen
wall development, and ROS generation or scavenging were
differentially expressed in the GMS mutant anther.
• compared to the WT anther these genes exhibit exhibit
opposite expression patterns in the GMS mutant anther,
which indicates that the hormone signals and energy
metabolism are disturbed or blocked during anther
development in the GMS mutant.
• These changes leads to male sterility.