This document summarizes research on leaf development and phyllotaxy (leaf arrangement) in plants. It discusses how KNOX proteins accumulate in the shoot apical meristem (SAM) but are excluded from leaves. Application of the plant hormone NPA arrests leaf initiation, showing KNOX proteins regulate this process. It also describes how the microRNA miR166 controls expression of the PHB gene involved in abaxial-adaxial patterning in leaves. Different leaf arrangements like alternate, opposite, whorled and spiral are defined. Experiments demonstrate the SAM maintains itself and periodically forms incipient leaf primordia in specific positions influenced by existing leaves. Evidence suggests the plant hormone auxin is a key signal regulating

1. LEAF DEVELOPMENT AND
PHYLLOTAXY IN PLANTS
CENTRAL UNIVERSITY OF KARNATAKA
Akash. V
MSc life sciences
Roll no. 2019MLS01
Department of Lifesciences

10. KNOX immunolocalization in maize shoots.
A. KNOX proteins accumulate in the SAM but are excluded from leaves and leaf founder
cells located in the periphery of the SAM (arrow in A).
B. On application of NPA, leaf initiation is arrested and there is no loss of .
KNOX accumulation in the SAM like A.
KNOX GENE INHIBITION

13. miR166 PHB mRNA
PHB-1D mRNA
In wild-type plants, miR166
binds to the PHB mRNA and
degrades it on the abaxial
side of the leaf primordium.
In phb-1d plants, base changes
in the PHB mRNA prevent
miR166 from binding to it,
allowing it to accumulate
throughout the leaf primordium.
Control of PHB expression by miRNA

17. PHYLLOTAXY
Distichous or alternate -- When each node has one organ, and those at adjacent
nodes are opposite, as in maize.
Decussate -- Two opposite organs at each node that are offset by 90° from those at
adjacent nodes. – Coleus. Tricussate- Nerium
Spiral -- a single organ at each node, but with 137 degree turn°.
Whorled -- used for more than two organs at each node, e.g. flowers- sepal & petals
Arrangement of leaves on stem

18. Cross section of rice
apex
Alternate Opposite Whorled

19. How primordia develops on Shoot apical meristem?
• The apical meristem is a pattern-generating machine.
• The WUS/CLV engine (yellow) maintains a supply (arrows) of cellular material to the rest of the
meristem (blue) to maintain its size and shape.
• At periodic intervals (termed plastochron), meristem tissue becomes determined to form a leaf
primordium at a site termed incipient primordium1 (l1; mauve).
• The position of l1 is determined by the positions of the previously formed leaf primordia, in this
example P1 and P2 (green).

20. P3
P2
P1
I1
I2
Two days later, the apex
was examined
a
P3
P2
P1
I1
I2
I1 (shown in black) was surgically
isolated from the rest of the
meristem, by cutting along the red
line.
P3
P2
P1
I1
I2
I3
Instead of emerging at its expected
position (star), I3 shifted towards I1.
This experiment shows that I1
influences I3 position.
Positions of I2 and I3; older leaves
have been cut away.

21. Is there any cross talk between primordia and SAM?
A single L1 cell of Betula SAM was injected at the
SAM centre with a fluorescent dye
Injection into the central zone shows that it is
symplastically isolated from the peripheral zone.
Dye does not move in PZ
Curved arrows show direction of dye movement.
Dye moves with in 5 sec.

22. Occasionally, some CZ
injection showed symplastic
connections with discrete
domains in the PZ. T
The positions of these
symplastic connections
appeared to be related
with position of incipient
primordia.

23. Wild-type Arabidopsis
Shoot apex
pin1 shoot
apex
What is the nature of Signal regulating primordia initiation?
Arabidopsis reduced polar auxin transport mutant Pin1 shows pin shaped inflorescence Evidences
favors auxin as regulatory molecule

24. 38 hours after application 4 days after application
Applying a spot of exogenous auxin (shown as a red blob)
stimulates outgrowth of primordium in the pin1 mutant.

25. What exactly happens in shoot apex? Where exactly auxin
moves?
A. In apex PIN1 protein was seen in epidermis and
vasculature of leaf primordia, also well L1 layer
and at lower levels in L2
B. A marked induction at the meristem flank (A)
corresponded with the site of incipient and
young leaf primordia.
C. PIN1 signal in the L1 and L2 layers was
consistently localized to the apical side of the
cells, that is, pointing to the meristem summit (C,
arrowheads), except for the cells close to the site
of incipient leaf initiation (I1). In these cells, the
signal pointed to I1 (Fig. 1c, arrows). At the
summit itself, the signal was more diffuse than at
the flank (C).

26. The Meristem Marker SHOOTMERISTEMLESS (STM) is downregulated as PIN1 is upregulated at Incipient Primordia
• At white arrowheads PIN1 is higher and STM is missing Auxin is trafficked apically through the epidermis and
becomes concentrated at a point on the meristem flank that corresponds to the next incipient primordium.
• The establishment of this auxin concentration peak is critical to the proper initiation of the primordium.
• Auxin is then drained through the center of the primordium

27. Auxin accumulation in young primordia. PIN1 is induced in
young primordia.
It becomes localized to the side of the cells that points to the
centre of the primordium (light blue).
This results in the accumulation of auxin in the primordium
and its withdrawal from the surrounding cells (bluearrows).
The resulting auxin gradient (red) confers positional
information to the cells allowing them to establish organ and
boundary identity.
Inset: Location of the P1 position in the context of the
apex.

28. I1
TIME
After primordium initiation, PIN1 distribution changes, directing
auxin flow into the developing midvein

29. Thank you