1. During prophase I, homologous chromosomes correctly recognise each other, pair and cross over
Homologues seen linked by Crossovers at Metaphase I
1 Diploid cell
Telophase II
Diplotene
Diakinesis
Pachytene
Zygotene
Leptotene
Metaphase I
Anaphase I
Telophase I
Sister chromatids
are separated during 2nd division
Homologues
separate
and segregated
4 Haploid cells
Meiosis

2. (Pairing homoeologous 1) controls correct pairing and recombination
Bread wheat-Triticum aestivum 2n= 6x= 42 1 2 3 4 5 6 7
Genome A
Genome B
Genome D
Homoeologues 1A 1B 1D
Ph1 locus
(Pairing homoeologous 1)
controls correct pairing and recombination
Homologues 1D

3. How Ph1 was first reported?
Both deleted chromosome 5B in wheat hybrids inducing crossovers between homoeologues
Sir Ralph Riley UK
Dr Ernie Sears US
The major regulator of pairing and recombination in wheat on 5B
Termed the locus, Ph1, (Pairing homoeologous 1), assuming it blocked homoeologous pairing, therefore recombination

4. What they saw in a wheat hybrid
Wheat-rye hybrid n = 28 1 2 4 3 5 6 7
Genome A
Genome B
Genome D
Rye
No homologues
Wheat-rye hybrid Ph1-
Wheat-rye hybrid Ph1+
Metaphase I
Up to one crossover
Up to 7 crossovers

5. Question 1. Does Ph1 actually block homoeologues pairing?
After 50 years the key antibodies to meiotic proteins available to answer two questions
Question 1. Does Ph1 actually block homoeologues pairing?
Early meiosis
Zygotene
Basics of chromosome pairing
Pachytene
homoeologues
“Glue added = amount of pairing

6. Ph1 has been named incorrectly for
In wheat-wild relatives hybrids, experimented on by Riley and Sears, there are no homologues, only homoeologues
Amount pairing= ”Glue” magenta
DNA-blue
Synaptonemal complex formed
“Pachytene”
Central Element
“Glue”
“Glue” or pairing (µm)
Pairing or Synapsis %
Wheat-Rye -
378.50
26%
Wheat-Rye +
403.95
27%
Homoeologue pairing is not reduced in wheat-rye hybrid by the presence of Ph1.
homoeologues
Wheat-rye hybrid Ph1-
Wheat-rye hybrid Ph1+
Ph1 has been named incorrectly for
50 years
Martin et al Nature Communications 2014

7. What happens in hexaploid wheat when homologues are present?
Wheat- rye hybrid 1 2 4 3 5 6 7
Genome A
Genome B
Genome D
Rye
When homologues are absent, Ph1 can’t block homoeologues pairing
Genome A
Genome B
Genome D
Wheat
Ph+
What happens in hexaploid wheat when homologues are present?
Ph-

8. The pairing of 15 homologous arms reveals that with Ph1 present:
1DL chromosomes only pair at telomere bouquet stage with Ph1 present
Homologous segments
Nature Cell Biol 2004
Ph1+
Ph1-
telomeres
In the start of meiosis, telomeres clustered (bouquet stage)
The pairing of 15 homologous arms reveals that with Ph1 present:
homologous arms pair correctly at the start of meiosis
Centromeres also pair correctly with Ph1 Nature 2001
So Ph1 promotes homologue pairing at the start of meiosis
(rather than block homoeologue pairing)

9. Question 2: When does Ph1 block recombination process between homoeologues?
Wheat- rye hybrid n = 28 1 2 4 3 5 6 7
Genome A
Genome B
Genome D
Rye
Wheat-rye hybrid Ph1+
Wheat-rye hybrid Ph1-
Up to one crossover
Up to 7 crossovers

10. = Ph1 blocks recombination between homoeologues- When? leptotene
ONE CHROMOSOME CUT
PAIR OF HOMOLOGUES or
HOMOEOLOGUES 5’ 3’
STRAND INVASION
DNA synthesis
EXPOSED SINGLE STRANDED 3’ END
Rad51
spo11
leptotene
DOUBLE HOLLIDAY JUNCTION 5’ 3’
diplotene
MLH1 protein
MLH1 site
crossover =
Alternative paths
Crossovers 5’ 3’
Non Crossover 5’ 3’
DNA STRANDS CUT AT ARROWS -
Metaphase I

11. 21 MLH1 sites whether Ph1 is present or absent in the hybrid
Wheat-rye Ph1-
7 crossovers- 7 MLH1 sites expected
MLH1- green
DAPI- blue
Wheat-rye Ph1+
one crossover- one MLH1 site expected
Martin et al Nature Communications 2014

12. Summary of data in hybrids
Homoeologues pair whether Ph1 is present or absent.
Recombination machinery loads with or without Ph1, but stalls with the loading of the last protein- MLH1.
Stalling is partially alleviated by deleting Ph1.
Ph1 affects the MLH1 complex and whether crossovers occur or not
Kinase
CDK2
MLH1
MLH3
EXO1
MLH1 complex

13. How to get 21 MLH1 sites = 21 crossovers??
Wheat hybrids Ph1-
21 MLH1 sites=7 crossovers
How to get 21 MLH1 sites = 21 crossovers??
It seems how well the wheat hybrid is growing……
CER + fertiliser
Glasshouse + fertiliser
Now ring bivalents
Rod bivalents
- fertiliser

14. The two Ph1 effects-promotion of homologue pairing in wheat and homoeologue recombination in hybrids defined to a kinase locus
Further deletions
Cdk-like genes
Kinases
(Griffiths et al Nature 2006)

15. Consistent with the presence of a kinase locus
Phosphorylation levels reduce with Ph1 present Greer et al Plant Cell 2012
Increasing phosphorylation levels mimics effect of deleting Ph1 Knight et al Funct Intreg Genomics 2010
Phosphorylation level
Ph1+
Ph1-
Phosphorylation of histone H1 during meiosis
Metaphase I
Wheat-rye hybrid treated with Okadaic acid-Rod bivalents

16. We have shown that: Ph1 maps to a kinase locus region –Nature 2006
Ph1 reduces meiotic phosphorylation levels-Plant Cell 2012
Ph1 promotes correct pairing of centromeres-Nature 2001
Ph1 promotes correct pairing of chromosome arms-Nature Cell Biol 2004
Ph1 regulates the conversion of MLH1 sites to crossovers- Nature Communs 2014
Increased phosphorylation levels mimic the effect of deleting Ph1 - Funct Intreg Genomics Plant Cell 2012
At increased phosphorylation levels, the ratio of MLH1 sites to crossovers is closer to 1:1 in healthy plants

17. A C elegans study (PLoS Genetics 2014) concluded:
reduced meiotic phosphorylation levels promote:
correct pairing of centromeres
correct pairing of chromosome arms
the conversion of MLH1 sites to crossovers
At increased phosphorylation levels, the ratio of MLH1 sites to crossovers is closer to 1:1 in healthier worms
Phosphorylation may be a universal regulator of correct meiotic prophase chromosome dynamics

18. Graham Moore- My thanks to…
Genomics- Comparative-BAC library-mutants-Tracie Foote, Michael Roberts, Lijia Qu, Terry Miller, Steve Reader, Simon Griffiths, Sebastien Allouis, Rebecca Sharp, Kath Mortimer, Emilie Knight, Nadia Al-Kaff, Vera Thole, Ruoyu Wen, Boulos Chalhoub, Shahryar Kianian, Dupont-Pioneer
Modelling-Faridoon Yousafzai, Nadia Al-Kaff, David Richards, , Azahara Martin, Martin Howard, Peter Shaw
Phosphoproteomics- advanced mass spec-Azahara Martin, Ali Pendle, Isabelle Colas, Alex Jones, Peter Shaw
Cell biology-Shahal Abbo, Luis Aragon, Fadri Martinez, Pilar Prieto, Mike Wanous, Isabelle Colas, Emma Greer, Azahara Martin, Danielle Monk, Lola Rey, Peter Shaw
Brachypodium/Arabidopsis- Ruoyu Wen, Ali Pendle,Vera Thole, Philippe Vain, John Doonan, Peter Shaw