1. Plant Reproduction
BIOL 304
11/21/ /26/2008

3. Transition to reproduction
Flower organ development
Gametogenesis and fertilization
Plant Reproduction

4. Transition to reproduction
Flower ?
Main inflorescence shoot in middle, that can branch into lateral shoot
At end of each shoot – place to make flower
Each inflorescence shoot can make leaves and lateral organs
What regulates transition from vegetative growth (main function to increase biomass of plant) to making plant sexually ready
Inflorescence
Vegetative phase
Reproductive phase

5. Production of flowers involves two transitions in Arabidopsis
Convert SAM to inflorescence meristem (infinite, making lateral organs)
2. Convert inflorescence meristem to floral meristem (terminal, flowers)
SAM convert to meristems that can make flowers
Influorescence mersitems can produce more lateral branches – can further branch into tertiary branches
Infinite branching as long as plant’s age allows – produces branches and leaf organs
Convert influorescence to floral mersitem – make final flower (cluster of flowers at each end of inflorescence shoot)
Once floral meristem finishes function to make flowers, some cell activity lost at that region – terminal organ
SC: stem cell
P: organ primordia
Se: sepal

6. Factors regulating the transitions
Vegetative meristem
Genes (flowering-time genes and floral identity genes)
Light (photoperiod)
The biological clock
Temperature
Hormones
Many factors regulate transition-
Vegetative mersitem can convert into inflorescence meristem
Inflorescence meristem
Floral meristem

7. Flowering-time genes
affecting the transition of vegetative growth to reproductive growth
Flowering-time genes regulate general transition from vegetative to reproductive growth
Emf1 and emf2 mutants flower during seedling stage – don’t have to go thru long process of making more shoot, root, etc.
Very early on, seedlings can make flowers – genes negative regulator of reproduction
(early flowering mutants – negative regulator of reproduction; promote vegetative growth – so under normal conditions, go to vegetative growth) WT
emf1
emf2
embryonic flower

8. Floral identity genes
affecting formation of inflorescence and floral meristems
Flower (from Floral meristem)
Floral identity genes – inflorescence and floral meristem affected
Flower from floral meristem and inflorescence from inflorescence meristem – how regulated?
Other types of mutants can tell function
Inflorescence (from Inflorescence meristem)

9. Mutations in floral identity genes
Terminal flower 1 gene normal function is positive regulator of inflorescence (when mutant, lose inflorescence shoot activity = flower early; SAM (vegetatuve) converted to floral mersitem (for making flowers))
Leafy – delay flowering; thus leafy gene acts as negative regulator of inflorescence meristem
terminal flower 1 (tfl1):
Convert the inflorescence meristem to the flower meristem.
leafy (lfy): produce more inflorescences,
delayed flowering

10. Factors regulating the transition to reproduction
Vegetative meristem
EMF
Inflorescence meristem
LFY
TFL
Floral meristem

11. The discovery of photoperiodism
Garner and Allard (1920’s)
Soybeans (Glycine max) planted over a three-month period all flowered about the same time

12. Many more experiments were followed:
Eliminate a variety of environmental conditions: Nutrition, temperature, and light intensity
Relative length of day and night decides the flowering time
Photoperiodism: ability of an organism to measure the proportion of daylight during a 24-hour period

13. Photoperiod
Varies according to the latitude and seasonal changes.

14. Critical daylength Critical Daylength (CD)
Xanthium: a short day plant, flowers when CD is LESS than 15.5 hours.
Hyoscyamus: a long day plant, flowers when CD is MORE than 11 hours.

15. Plants are induced to flower by different photoperiods
short day (SD) : plants are stimulated to flower when the length of day falls below a threshold
long day (LD): plants are stimulated to flower when the length of day exceeds a threshold
Day neutral (DN): plants flower indifference to the changes of day length.
Long-short-day: flowering requires certain number of short days are preceded by a certain number of long days.
Short-long-day: flowering requires certain number of long days are preceded by a certain number of short days.
Intermediate-daylength: not flowering if the daylength is too short or too long.

16. Do plants really measure the length of the daylength?

17. Hamner and Bonner (1938): Xanthium strumarium, a SD plant with CD = 15
Hamner and Bonner (1938): Xanthium strumarium, a SD plant with CD = 15.5 hours
Xanthium flowers when the dark period exceeds 8.5 hours.
Short interruption of dark period, even by a pulse of light as short as 1 minute delays flowering.
The relative length of dark is not the determining factor.

18. Similar results were obtained with other SD plants.
For LD plants
A longer dark period inhibits flowering.
Light break induces flowering.
SD: A longer dark period promotes flowering.
Light break inhibits flowering.

19. What tissues/organs perceive photoperiod?

20. Exp. 1: The leaf or apex of Perilla (a short day plant) was exposed to different daylength.
When leaf part treated w/ short day (regardless w/ what apex treated) – plants made flower
When only treated single leaf w/ short day – plant couldn’t make flower
The leaf, not the apex perceives photoperiod

21. Exp. 2: Grafting experiment with Perilla
Only single leaf from one of the plant treated – all 5 plants can make flower
Something made in leaf can be transferred
Flowering signal is grafting transmittable

22. The flowering signal: florigen
vegetative or reproductive growth?
the flowering signal is generated in the leaf
the signal goes one way: from the leaf to the apex
Grafting transmittable
SAM
Florigen can be transmitted to other plants thru grafting
Flower locus T protein claimed to be florigen – but some disputes
If you find signal or molecule that can greatly change flowering time – agriculture revolutionized
Florigen ?
Florigen
Florigen

23. The biological clock
Present in plants, animals, fungi, and some photosynthetic bacteria
An internal time measuring system (“clock”) that runs on its own with a periodicity of nearly 24 hours. It can be “reset” by external signals.
Temperature
Present in many different organisms
Very important to control daily activities of organisms
Movement of leaves, when to make flowers, etc.
Central component of clock is central oscillator – internal time measuring machine in many organisms – can run independently of external factors (intrinsic) – can also be regulated by external factors such as light and temperature
Output of clock represented in rhythmic activity – circadian

24. The Arabidopsis biological clock
The central oscillator: CCA1, LHY, and TOC1 (these are transcription factors) and other proteins
Three TF regulate expression of each other
-TOC1 is activator of CCA1 and LHY
-CCA1 and LHY are repressors that inhibit TOC1
In morning, TOC1 accumulate to certain abundant level – w/ more TAC1, other two genes expressed in morning
More expression of these two genes, more of those two proteins made – those two proteins bind to each other to form heteodimer – bind to promoter of TOC1 gene to inhibit its expression
When have more LHY and CCA1 = less TOC1 protein
At end of day, TOC1 protein eventually disappear thru protein degradation and lack of making new TOC1 protein, so at end, lack of TOC1 will make expression of these two genes turned off – so no more LHY and CCA1 protein made
In evening, no more genes to expression this point, eventually LHY and CC1 protein disappear – as a result, TOC1 no longer being de-repressed – so expressed and accumulate in evening
In morning, CCA1 and LHY more thus TOC1 once again repressed

25. The Arabidopsis biological clock
CCA1 and LHY are expressed during the day and together repress expression of TOC1 during the day
These three make up central component – expressed in circadian manner
Expression of CCA1 and LHY and TOC1 is circadian regulated
Arabidopsis seedling expresses CCA1::luciferase
TOC1 is expressed at night and is required for activation of CCA1 and LHY1, beginning just before morning

26. Mutations in the clock genes
Lack of the nyctinastic movement: diurnal rise and fall of leaves
Altered flowering time in some mutants
cca1: early flowering
lhy: early flowering
toc1: early flowering
Some other clock mutants can be late flowering
Different type of mutations – some make clock go fast and others make clock go slower

27. Temperature: Vernalization
Vernalization: low temperature treatment can promote flowering in some plants.
The vernalization-effective temperature and duration of low temperature treatment may vary.
Vernalization is perceived by the shoot apex.
The vernalization state is grafting transmissible.
Vernalization—low temperature treatment can promote flowering time in some plants
Low temperature treatment at earlier time but can still affect later plant development for plants to make flowers or not
Affective temperature can vary and in different plants
Some plants don’t need vernalization - ex. plants grown in cold regions – have to go thru winter time so have to be vernalized – in high temp, don’t need vernalization
Perceived by shoot apex – if treat dry seeds w cold temp – don’t matter – only when seed germinates, shoot apex active and can perceive cold temperature and affect flowering
Not known which molecule induced – induced stage can stay and later on affect flowering
Also grafting transmissible

28. Cold acclimation
Cold acclimation is different – couple of days of cold weather before deep winter – very important for plant b/c repeated cold treatment to trees can dramatic increase plant tolerance to freezing temps later on in deep winter time
Cold acclimation can be induced quickly (not long term like vernalization) and it doesn’t affect flowering time
Can be induced quickly
Increases plant resistance to freezing stress
Does not affect flowering time.

29. Hormone GA regulates flowering time
GA1: an enzyme involved in GA biosynthesis
ga1: In addition to the dwarf phenotype, the mutant flowers late under LD conditions and does not flower under SD conditions.
GA treatment promotes flowering time.
GA positively regulates flowering time

30. Inflorescence meristem
Flower development in Arabidopsis
Vegetative meristem
Inflorescence meristem
Transition to reproduction:
Genes & other factors
Flower organ development stage less regulated by environmental factors (light, temp) , but more by intrinsic - genes (organ identity genes)
Floral meristem
Flower:
sepals, petals, stamens,
and carpels
Flower organ development:
Organ identity genes

31. Flower organs petal stamen carpel sepal Petals - attract polinators
Stamen – filament and anther (pollen sac)
Carpel – stigma (flower perceives pollen; pollens land) and style – long or short neck – connects stigma to ovary; Ovary houses ovules – contain egg cells and central nuclei for double fertilization
Sepal – green leaf like structure; degenerative leaves – can perform limited photosynthesis – protect flower bud before it opens
Receptacle – holds the flower

32. The flower is generated from the floral meristem
Floral meristem can give rise to cluster of flowers, but in some one flower meristem can only give rise to one flowers (ex. Roses)
the floral meristem

33. Produced in 4 concentric whorls with the same order
Flower organs
Produced in 4 concentric whorls with the same order
sepal (whorl 1) stamen (whorl 3)
petal (whorl 2) carpel (whorl 4)
w/ Arabiposis flowers – many mutants identified to understand processes of flower organ formation
Four whorls –
3rd whorl – 6 stamens
Middle – two carpels

34. sepal-petal-stamen-carpel stamen-carpel-stamen-carpel
Mutants have weird flower structure – puzzle for many years
Apetala2-2 mutant = only see stamen, carpel, stamen, etc.
Ap1 mutant is similar
sepal-petal-stamen-carpel
stamen-carpel-stamen-carpel
(the ap1 mutant is similar)

35. Aptala3 and Pistillata = see sepal, sepal, carpel, carpel (lose petal and stamen; 2nd and 3rd whorl are lost) wt
apetala3 (ap3)
pistillata (pi)
sepal-petal-stamen-carpel
sepal-sepal-carpel-carpel

36. sepal-petal-stamen-carpel sepal-petal-petal-sepal
Agamous 1 – see sepal and petal only; so lost two inner whorls
sepal-petal-stamen-carpel
sepal-petal-petal-sepal

37. The “ABC” model for flower development
AP1, AP2 B
AP3, PI C AG
ABC model used to explain weird flower structure
A, B, and C genes function to specify different structures in diff. whorls
A+B = 2nd, B+C = stamen, and C alone = carpel = only when functioning in all three categories, have normal flower
A and C genes are antagonizing each other in their function – only in certain physical region
The ABC genes function in the distinct fields.
The A and C genes are mutually exclusive in their expression.

38. WT
Ap1 and ap2 mutants are A genes – if knock them out, C will expand to A territory – don’t have sepal and petal, but will have carpel in sepal position b/c C expanded into A region and C only specifies carpel
So, get carpel-stamen-stamen-carpel
ap1 or ap2
The A genes: ap1 or ap2 mutants should (and do) make carpel-stamen-stamen-carpel

39. WT
Ap3 and pi are B genes, if lost them, lost combination of AB and BC, so lose petal and stamen and have sepal-sepal-carpel-carpel
ap3 or pi
The B genes: ap3 or pi mutants should (and do) make sepal-sepal-carpel-carpel

40. The C genes: ag mutants should (and do) make sepal-petal-petal-sepalWT
If don’t have C genes, loss of C, make A expand to C’s territory and lose stamen and carpel
So have sepal-petal-petal-sepal
ABC genes are all transcription factors that act as master regulators to control downstream genes – all together to regulate floral organ formation
All ABC genes function within physical restriction space
The ABC genes are TF that set up the boundary of gene expression to determine which organ will be formed, and where
Additional downstream genes are required to ensure the property development of each organ ag
The C genes: ag mutants should (and do) make sepal-petal-petal-sepal

41. Flower organ function:
Gametogenesis and Fertilization
Gametogenesis involves 2 meiosis – takes place in two different locations

42. Male gametogenesis
Diploid pollen mother cells undergo meiosis to produce a tetrad of haploid microspores.
Each microspore develops into a pollen grain containing two haploid cells (mitosis I):
Pollen mother cells undergo meiosis to produce tetrad of haploid microspores – attach to each other
Later separate into individual pollen cells – undergo mitosis to give rise to two cells for each pollen cell
One cell engulfes the other cell (small = generative cell – lives inside the large, vegatative cell)
the generative cell (small)
The vegetative cell (large)
generative
cell

43. Vegetative cell grows to produce pollen tube
Goal of pollen tube is to deliver male sex cells to female
Generative cell produce 2 sperm cells via mitosis II – 2 sex cells for double fertilization
the vegetative cell grows to produce the pollen tube
the generative cell produce 2 sperm cells (mitosis II)

44. Female gametogenesis
an ovule primordium emerges
as a bump on the inner wall
(placenta) of the ovary
the megasporocyte
undergoes meiosis to produce
4 haploid cells, only one of
which (the megaspore)
survives.
Ovule inner wall can bud to form ovule primordium – one of the cells undergo meiosis to produce 4 cells – only one survives
Further divides three rounds to make 8 cells

45. Female gametogenesis placental wall
the megaspore undergoes 3 mitotic divisions to produce 8 cells:
3 antipodal cells
2 synergid cells
2 central cell nuclei
1 egg cell (EC)
3 antipodal cells (die eventually)
2 synergid cells located right at entrance of ovule – micropyle
Pollen tube navigate all the way to micropyle and deliver sperm cell
In end, synergid cells die – only 3 cells survive

46. Female gametogenesis
placental wall

47. Double fertilization
Pollens land on the stigma, hydrate, and begin to germinate the pollen tube

48. Pollen tubes grow, by tip growth, down through the stigma and style and into the ovary, toward the ovules.
The pollen tube navigates to the micropyle and discharges the two sperm cells.
Pollen tube doesn’t penetrate but navigates in b/w of cells

49. Double fertilization
One sperm fertilizes the egg cell to develop into the embryo.
the other sperm fertilizes the diploid central cell nucleus to develop into the endosperm.
Ovule
Antipodal cells
Central cell nuclei
One sperm cell will fuse w/ egg – into embryo and other will fuse w/ central nuclei – into endosperm
Egg
Synergids
Pollen tube
Micropyle
Sperms

50. Plant reproduction
Ovule (1 to many)
Ovary
Silique

51. Fruit development The ovary and other tissue together produce a fruit.
Fruit is important for seed dispersal in many species
Many foods are also called “vegetables”: tomatoes, pea pods, squash
Fruit size, texture, and sugar content are determined by genes.
Ethylene stimulates fruit ripening.

52. Life cycle of a flowering plant