The document discusses leaf growth and differentiation, outlining the stages of leaf ontogeny from initiation at the shoot apical meristem (SAM) to the completion of histogenesis. It details the role of genetic factors, including the plastochron gene in rice and various homeobox genes, in regulating leaf formation and development. The mechanisms of auxin transport and gene interactions that influence leaf primordium initiation are highlighted, emphasizing the complex regulatory networks involved in leaf morphology.

1. Leaf Growth and
Differentiation
Dr. Renu Jangid

2. Leaf Initiation
• It is often convenient to divide leaf ontogeny into three phases
depending on the time at which different leaf features become
determined. During the first phase, the leaf primordium is initiated
from the SAM and acquires its identity.
• During the second phase, the major parts of the leaf become
determined and the leaf gets its basic shape, and in the third phase, leaf
histogenesis is completed.

3. • Leaf primordia are stated to come into being following a change in the
orientation of cell enlargement and division in both the tunica and corpus of
the flank meristem of SAM.
• This change results in the formation of a protrusion from SAM, called leaf
buttress (Fig. 5.3), resulting in asymmetric or symmetric enlargement of
SAM depending on alternate or opposite phyllotaxy. The interval between
the initiation of two successive leaf primordia (or pairs of leaf primordia in
opposite phyllotaxy) is called a plastochron, and the changes in the SAM
during this interval are called plastochronic changes.

5. • A PLASTOCHRON1 ( PLA1 ) gene has been known from rice plant, and this
gene is implicated in the regulation of duration of vegetative phase by controlling
the plastochron intervals.
• The leaf primordia of plants start from SAM, which is an embryonal cell
population with a “tunica-corpus” structure. The structure can be divided
into three layers. The tunica cells divide into L1 and L2, and the corpus
cells divide into L3.

6. Leaf Determination and Development
• In median longitudinal sections of shoot apices of most plants examined, the first
sign of leaf initiation is seen as localized periclinal cell divisions at some distance
from the apical meristem.
• The sites of cell division activity in the shoot apical meristem presaging leaf
primordia are not randomly selected but are arranged according to a genetically
determined pattern or phyllotaxy, characteristic of each species.
• In monocots, cell divisions are confined to the outer tunica layer, whereas in
dicots, cells in the first and second, or second and third, or even deeper layers of
the apical meristem may show accelerated mitotic activity.

7. • In the young primordium, the centrally located group of cells in the original bump
functions as the apical meristem of the leaf; cell divisions begin at its margins or
in the region between the midrib and margins.
• This latter growth continues for a longer period than the apical growth, and it too
ceases after producing a dorsoventrally symmetrical leaf blade or lamina.
• The remaining period of growth of the leaf is completed by cell enlargement. The
lower part of the leaf blade extends into a stalk or petiole, and many leaves
develop at the base of the petiole, small scalelike or leaflike appendages called
stipules.

8. • The framework within which the three layers of the shoot apical meristem
contribute tissues of the mature leaf has generally assigned the L1 layer to
the formation of the epidermis, the L2 layer to the formation of the upper
palisade and the lower spongy mesophyll, and the L3 layer to the middle
mesophyll and the vascular tissues.
• In most monocots and some dicots, the base of the leaf blade is expanded
into a sheath, which clasps around the stem. Indications of the sheath are
seen in the tendency of the young leaf primordium to encircle the shoot
apex by lateral extensions.

9. • A critical review of leaf development in angiosperms reveals that there
are two classical mechanisms on how leaves develop:
• (1) The final shape of leaf with its fully differentiated tissues is
determined by a series of properly oriented cell divisions closely
accompanied by appropriate patterns of cell elongation.
• (2) Once apical growth of leaf primordium (leaf axis) stops, the
dorsiventral lamina is produced by specific marginal meristems and
submarginal meristems along the lateral sides of leaf axis (Fig. 5.4 ).

11. Genetic Control of Leaf Development
• Phyllotaxy is considered generally to be a non-mutagenic trait. Many
regulatory genes that function in leaf formation and positioning are known,
but intensive genetic screening has mostly failed to yield mutants that
specifically affect phyllotaxis. In other words, there is so far no homeotic
transformation of one phyllotaxy type into another type.
• Till now, information on the functions of several individual genes in the
control of leaf initiation and development has accumulated.

12. • The shoot apical meristem (SAM) comprised by the central zone (CZ), the
peripheral zone (PZ), and the organizing center (OC). At the apex of the
SAM, there is a cluster of slowly dividing cells, constituting the central
region of SAM. These cells are larger, possess stem cell functions, and play
a pivotal role in maintaining the meristem’s integrity.
• The rate of cell proliferation and growth in this central region often differs
significantly from that at the periphery. In the periphery of the central SAM
region, cell division rates are notably accelerated. These rapidly dividing
cells form the peripheral region of SAM, which serves as the origin for
organ primordia such as leaf primordia.

13. • Below SAM lies the organizing center, also known as the Rib Meristem
(RM). Within the SAM, the homeodomain transcription factor WUSCHEL
(WUS) is expressed in the organizing center (OC) to uphold stem cells in
the central zone (CZ). The migration of WUS to the central zone activates
the accumulation level of the CLAVATA3 (CLV3). CLV3 acts as a negative
regulator by encoding a secreted peptide. This peptide triggers the
transmembrane receptor kinase CLV1 in the organizing center, resulting in
the inhibition of WUS expression (Sassi and Vernoux, 2013).

14. • During the formation of plant leaf primordium, the plant hormone auxin is
the growth regulator of organ initiation. The highest local auxin
concentration observed in the L1 cambium of the SAM. This localized
increase in auxin concentration is facilitated by polarly localized PIN-
FORMED1 (PIN1) efflux transporters. The rise in auxin levels coincides
with the onset of leaf primordium formation, and the cellular response to
auxin is mediated by AUXIN RESPONSE TRANSCRIPTION FACTORs
(ARFs). Auxin level has a negative effect on SAM size.

16. • SAM is divided into three functional regions [central region (CZ),
peripheral region (PZ) and costal region (OC)] and Layer1 (L1), Layer2
(L2) and Layer3(L3). WUS activates CLV3, and CLV3 further binds to
CLV1/2, thereby inhibiting WUS expression. Auxin accumulation in the
flanks of SAM through PIN1/AUX1 mediated polar transport triggers
primordium development. In addition, KNOX1 maintains the role of stem
cells, positively regulates CK, negatively regulates GA signaling through
IPT7 and GA20ox, and ARF regulates the emergence of young primordia.

17. • Among genes that control leaf initiation, most are known in Arabidopsis and
maize. The class 1 KNOX members ( SHOOT APICAL MERISTEMLESS ; STM
; AT1G62360 ; KNAT1 , KNAT2 ; AT1G70510 ; and KNAT6 ; AT1G23380 ) are
key factors in the formation and maintenance.
• Studies in Arabidopsis thaliana, Zea mays L., and Antirrhinum majus have
unveiled two molecular mechanisms governing the initiation of leaf primordia
(Barkoulas et al., 2007; Yan et al., 2008; Barton, 2010). The first mechanism
involves the polar localization of the auxin transporter PIN1, ensuring the
transport of auxin to the initial site of leaf primordium. Additionally, accumulated
auxin inhibits the expression of the Class I KNOTTED1-like homeobox (KNOX1)
gene.

18. • The second mechanism is the mutual inhibition of the tip meristem maintenance
gene KNOX1 and ARP [ASYMMETRIC LEAVES1(AS1)/ROUGH
SHEATH2(RS2)/PHANTASTICA (ARP)].
• The downregulation of KNOTTED1 ( KN1 ) class of homeobox-containing plant
genes that were originally identified in maize in fact provided an early molecular
marker of leaf initiation in SAM (Brutnel and Langdale 1998 ; Sinha 1999 ).