ORGANOGENESI: DIFFERENZIAMENTO DEI PRIMORDI FOGLIARI

Le foglie si formano nello sviluppo post-embrionale Cotyledons (embryonic leaves) Embryo inside seed zygote Germination Embryonic development

Post-embryonic development True leaves Cotyledons (embryonic leaves) Post-embryonic development

Sono formate dal meristema apicale del germoglio (SAM) SHOOT APICAL MERISTEM Leaf formation True leaves zygote Germination

Il SAM si forma durante l’embriogenesi Shoot apical meristem Apical Basal TOP DOWN Shoot apical meristem Laux, T, Jurgens, G. (1997) Plant Cell 9: 989-1000

Dopo la germinazione il SAM forma le foglie Leaves Shoot apex at germination Post-embryonic leaf formation Cotyledons Leaves Cotyledons Reprinted by permission from Macmillan Publishers, Ltd: NATURE. Long, J.A., et al., 379: 66-69, copyright 1996.

diversi stadi nello sviluppo dei primordi Organogenesi: poche cellule negli strati L1 e L2 nella zona periferica acquistano l’identità di iniziali (founder cells) della foglia. Cominciano a dividersi più rapidamente delle cellule circostanti e formano una zona distinta dal resto del doma (primordio). Sviluppo di differenti regioni nella foglia: regioni dei primordi acquisiscono l’identità delle diverse parti della foglia. Tre assi di sviluppo: adaxiale/abaxiale; prossimale/distale; mediale/laterale Differenziamento di cellule e tessuti: Con la crescita della foglia cellule e tessuti si differenziano: L1 epidermide; L2 mesofillo; L3 elementi vascolari e cellule della guaina del fascio

DIVISIONI CELLULARI NELLA FORMAZIONE DEL PRIMORDIO (arabidopsis) divisioni periclinali nello strato più interno della tunica divisioni periclinali anche negli strati meno interni della tunica e divisioni meno orientate divisioni anticlinali nelllo strato esterno della tunica per formare il protoderma

Le foglie in formazione hanno una loro polarità intrinseca Peripheral Central Leaf Cot Cot Leaf

Assi di asimmetria nella foglia

La polarità è evidente fin dagli stadi iniziali The adaxial side is towards the center of the plant Peripheral Central Adaxial Abaxial One way to remember which word is which – ad is like adjacent (from Latin adjacere to lie by the side of), ab is like abduct – to take away (from Latin abducere, abductum to lead away).

In quale posizione si formano sul germoglio? (fillotassi) Alternate Opposite Whorled Spiral

Alternata One leaf at a time, 180° apart, as in rice or other grasses. TEM of rice apex Cross section of rice apex One leaf at a time, 180° apart, as in rice or other grasses. Candela, H. et al. (2008) Plant Cell 20: 2073-2087; Itoh, J.-I., et al. (2000) Plant Cell 12:2161-2174

Opposta Two at a time, 180° apart at each node. Sometimes pairs alternate by 90° at successive nodes.

Verticillata Three or more leaves at each node, as in the horsetail (Equisetum). This whorled arrangement of organs is also common in floral organs such as petals and sepals. Photos courtesy of tom donald

Spiralata In most plants, such as this succulent, leaves form in a regular spiral pattern. Photos courtesy of tom donald

spiralata In plants with spiral phyllotaxy, leaves form at about 137° apart. 137°

Spiral phyllotaxy

Spiral phyllotaxy

Spiral phyllotaxy A line through sequential leaves makes a spiral.

Spiral phyllotaxy The NEXT leaf to form is called the Incipient primordium (I1). I1

Spiral phyllotaxy The one that will form after that is called I2….etc.

Fillotassi spiralata in apice di tabacco Poethig , R.S. and Sussex ,I.M. (1985) The developmental morphology and growth dynamics of the tobacco leaf. Planta 165: 158-169. Copyright (1985) Planta. Reprinted with kind permission of Springer Science+Business Media.

Cosa determina la posizione del primordio incipiente? Surgical experiments demonstrate that leaf placement is determined by pre-existing primordia Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

I2 This tomato apex shows the positions of several primordia (P) and incipient primordia (I). Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

The expected position for I3 (*) can be found by tracing the spiral. This tomato apex shows the positions of several primordia (P) and incipient primordia (I). The expected position for I3 (*) can be found by tracing the spiral. Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

P3 P2 P1 I1 I2 I1 (shown in black) was surgically isolated from the rest of the meristem, by cutting along the red line. Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

Two days later, the apex was examined. Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

This experiment shows that I1 influences I3 position. Instead of emerging at its expected position (star), I3 shifted towards I1. This experiment shows that I1 influences I3 position. Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

Positions of I2 and I3; older leaves have been cut away. This scanning electron micrograph (SEM) shows that I3 formed approximately 180° away from I2. Note that the older primordia havae grown and are removed to better show the positions of I2 and I3. Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

The older primordia control the placement of the incipient primordia. What kind of signal or information is involved?

L’auxina è coinvolta nella formazione del primordio Wild-type Arabidopsis shoot apex. The meristem is covered by the leaves it has produced. Reprinted from Current Opinion in Plant Biology, 8 (1), Byrne, M.E., Networks in leaf development , 59-66, Copyright (2005), with permission from Elsevier .

L’auxina è coinvolta nel determinare la posizione del primordio meristem The apex of the pin1 mutant is bare – it fails to produce lateral organs. pin1 shoot apex Wild-type Arabidopsis shoot apex Reprinted from Current Opinion in Plant Biology, 8 (1), Byrne, M.E., Networks in leaf development , 59-66, Copyright (2005), with permission from Elsevier ; Reinhardt D et al., (2000) Plant Cell 12: 507- 518

IAA N H O OH The pin1 mutant is defective in the transport of auxin.

Auxin transport Auxin (IAA) is a weak acid. At the low pH of cell walls, it is protonated and uncharged (IAAH), allowing it to move through the plasma membrane. pH 5 pH 7 Cytoplasm IAAH Cell wall

Auxin transport In the less acidic cytoplasm, it loses the proton, becomes charged (IAA-), and cannot exit the call by diffusion through the plasma membrane. pH 5 pH 7 Cytoplasm IAAH IAA- Cell wall

Auxin transport The PIN1 protein is an auxin efflux carrier, transporting charged auxin back out of the cytoplasm. pH 5 pH 7 IAA- PIN1 protein IAAH IAA- Auxin efflux through PIN1

Auxin transport Net flow of auxin The subcellular localization of PIN proteins can be polar and coordinated between cells, causing directed auxin transport. In this diagram, the accumulation of PIN1 to the right of each cell causes a net flow of auxin towards the right. IAA- pH 7 pH 5 IAAH Net flow of auxin

Un massimo localizzato di auxina è richiesto per l’organogenesi Applying a spot of exogenous auxin (shown as a red blob) stimulates outgrowth of primordium in the pin1 mutant. Reinhardt D et al., (2000) Plant Cell 12: 507- 518

38 hours after application 4 days after application Reinhardt D et al., (2000) Plant Cell 12: 507- 518

Conclusion - Auxin transport and a local auxin maximum contribute to organ initiation. This conclusion is supported by imaging PIN1 distribution in living plants. IAA- pH 7 pH 5 IAAH

Visualizing PIN1 localization Emitted light PIN1 GFP GFP Excitatory light Green fluorescent protein (GFP) emits green light when excited by blue light. A protein’s position within a cell can be determined by making a fusion protein of it with GFP, and then looking for GFP fluorescence.

Visualizing PIN1 localization Reporter gene in the nucleus PIN1pro PIN1 GFP mRNA GFP PIN1 Translation Fusion protein PIN1 GFP Insertion into membrane

Visualizing PIN1 localization Using a confocal laser scanning microscope, PIN1:GFP protein distribution can be imaged in the shoot apical meristem. In this image, the green lines show the position of PIN1:GFP at cell membranes. PIN1 GFP PIN1pro PIN1 GFP mRNA Reproduced with permission - Development Gordon, S.P., Heisler, M.G., Reddy, G.V., Ohno, C., Das, P., Meyerowitz, E.M. Development, 2007, 134 (19): 3539-3548.

La distribuzione di PIN1 è dinamica durante l’organogenesi Plants expressing PIN1:GFP were examined over time to investigate the contributions of auxin flux to organogenesis. PIN1:GFP Positions of primoridia and incipient primordia

The orientation of PIN1 within cells is shown by white arrows, and indicates auxin flow.

I1 Auxin accumulates at I1 position Reprinted from Current Biology 15: Heisler, M.G., Ohno, C., Das, P., Sieber, P., Reddy, G.V., Long, J.A., and Meyerowitz, E.M. Patterns of auxin transport and gene expression during primordium development revealed by live imaging of the Arabidopsis inflorescence meristem,1899-1911, Copyright (2005), with permission from Elsevier.

La polarità delle proteine PIN 1 determina un massimo di auxina in I1

I1 This observation is consistent with the emergence of a primordium at the site of auxin application

dopo l’inizio della formazione del primordio la distribuzione di PIN1 cambia e orienta il flusso di auxina nel tessuto vascolare I1 P1 P1 I1 TIME Poethig , R.S. and Sussex ,I.M. (1985) The developmental morphology and growth dynamics of the tobacco leaf. Planta 165: 158-169. Figure 3 Copyright (1985) Planta. Reprinted with kind permission of Springer Science+Business Media. Adapted by permission from Macmillan Publishers, Ltd: Nature Reinhardt D., Pesce, E.-R., Stieger, P., Mandel, T., Baltensperger, K., Bennett, M., Trass, J., Friml, J., Kuhlemeier, C. Regulation of phyllotaxis by polar auxin transport. Nature 426, 255-260; copyright (2003).

Una successiva inversione nella polarità di PIN1 cambia la posizione del picco di auxina e specifica la posizione del nuovo primordio P3 P2 P1 I1 I2 time

Summary Organ initiation at the shoot apical meristem is determined by auxin distribution and PIN1 An auxin maximum is necessary and sufficient to specify the site of primordium formation Primordia affect auxin distribution and so placement of incipient primordia Auxin has been proposed to act as a morphogen – a generator of form

Come viene acquisita l’identità di foglia?

The meristem is a population of small, undifferentiated, dividing cells. A leaf primordium is a population of small, undifferentiated, dividing cells.

The differ in their expression of critical regulatory genes; the meristem expresses meristem-specific genes, and the leaf primordium expresses primordium-specific genes.

Ruolo dei fattori di trascrizione Poethig , R.S. and Sussex ,I.M. (1985) The developmental morphology and growth dynamics of the tobacco leaf. Planta 165: 158-169. Figure 3 Copyright (1985) Planta. Reprinted with kind permission of Springer Science+Business Media.

Geni di identità meristematica KNOX-1 Class I KNOX genes (KNOX-1) (KNOX means Knotted-like homeobox) Expressed in meristem Not expressed in incipient primordia Help maintain indeterminate growth

Class I KNOX genes KNOX genes are Knotted-like homeobox genes that encode homeodomain transcription factors. Genes described further are highlighted in yellow. Hao Yu, H., Yang, S.H., and Goh, C. J. (2000) DOH1, a class 1 knox gene, Is required for maintenance of the basic plant architecture and floral transition in orchid. Plant Cell 12: 2143-2160.

Espressione di KNOTTED KNOTTED (a KNOX-1 gene) mRNA accumulates in the meristem but not the leaf primordia (arrows) of Zea mays. Expression (purple stain) of KNOTTED mRNA in the meristem of Zea mays. Note the lack of expression in the leaf primordia (arrows). Down-regulation of KNOX-1 genes (that promote indeterminacy) contributes to the determinate growth pattern of most leaves. Jackson, D., Veit, B., and Hake, S. (1994) Expression of maize KNOTTED1 related homeobox genes in the shoot apical meristem predicts patterns of morphogenesis in the vegetative shoot. Development 120: 405–413. Reproduced with permission.

STM un gene di classe KNOX1 è necessario per la formazione del meristema The shootmeristemless mutant (stm) fails to form a shoot apical meristem during embryogenesis; notice the absence of leaf formation. Wild-type plant showing leaf formation at the shoot apex Reprinted by permission from Macmillan Publishers, Ltd: NATURE. Long, J.A., et al., 379: 66-69, copyright 1996.

Geni Primordio-specifici ARP ARP genes “ARP” is derived from three genes, ASYMMETRIC LEAF1, ROUGH SHEATH2, and PHANTASTICA ARP genes encode MYB transcription factors Expressed in cells of leaf primordia Promote determinate growth and differentiation

ASYMMETRIC LEAF1 (AS1) mRNA is expressed in cotyledons but not in the meristem . ARP Reprinted by permission from Macmillan Publishers, Ltd: NATURE. Byrne, M.E., et al., 408: 967-971. Copyright 2000..

In the stm mutant, AS1 is expressed in the meristem (arrow). ARP Wild type stm mutant Reprinted by permission from Macmillan Publishers, Ltd: NATURE. Byrne, M.E., et al., 408: 967-971. Copyright 2000..

L’espressione dei geni KNOX nel meristema reprime quella dei geni ARP

L’espressione dei geni ARP reprime quella dei geni KNOX

La sovraespressione di KNOX-1aumenta la complessità della foglia e la sua indeterminazione Arabidopsis Tobacco Maize WT OX WT OX WT OX Chuck G et al., (1996) Plant Cell 8: 1277-1289. Reprinted by permission from Macmillan Publishers, Ltd: NATURE GENETICS 31: 121 – 122. Hake, S., and Ori, N. Plant morphogenesis and KNOX genes. Copyright (2002).

Mutazioni loss of funcion arp hanno fenotipo simile alla sovraespressione di KNOX-1 Arabidopsis Maize Wild-type as1 WT as1 rs2 WT rs2 WT rs2 Reprinted by permission from Macmillan Publishers, Ltd: NATURE 408: 967-971. Byrne, M.E., Barley, R., Curtis, M., Arroyo, J.M., Dunham, M., Hudson, A., and Martienssen, R.A. Asymmetric leaves1 mediates leaf patterning and stem cell function in Arabidopsis. Copyright (2000). Reproduced with permission Development Schneeberger, R., Tsiantis, M., Freeling, M., Langdale, J. Development (1998) 125: 2857-2865.

Nei primordi i geni ARP agiscono come repressori trascrizionali dei geni di classe KNOX ARP heterodimer KNOX gene Guo, M., et al. (2008) Plant Cell 20:48-58

Geni di confine (boundary) sono necessari per la separazione degli organi Boundary genes Ensure a sharp boundary between leaf and meristem Expressed at organ boundaries Loss-of-function leads to “jagged” or fused organs

GENI CUC

Loss-of-function phenotype JAGGED LATERAL ORGAN (JLO) (LOBD gene family) JLO expression Loss-of-function phenotype JAGGED LATERAL ORGANS (JLO) is a boundary gene. Loss-of-JLO function causes fused or jagged organs. JLO coordinates KNOX-1 and PIN activities.

Summary A leaf acquires identity by turning OFF meristem genes and turning ON leaf genes KNOX-1, ARP and boundary genes encode transcriptional regulators that control expression of other genes Precise control of cell fates involves tight control of transcription by developmentally regulated activators and repressors

COME VIENE ACQUISITA LA POLARITA’?

Polarità anatomica e funzionale Abaxial surface - transpirational water loss, respiratory gas exchange Adaxial surface – light harvesting Most leaves have polarity – they are functionally and anatomically different on their upper and lower surfaces O2 CO2 Juarez, M. T., Twigg, R.W., and Timmermans, M.C.P. (2004) Development 131:4533-4544. Reproduced with permission.

Leaf Peripheral Central Leaves have an inherent polarity because one side is more central and one more peripheral.

The central side is adaxial, and peripheral is abaxial. Leaf Peripheral Central Adaxial Abaxial The central side is adaxial, and peripheral is abaxial. How does a leaf “know” which side is central?

The Sussex signal In the 1950s, Ian Sussex showed that a signal from the meristem is required for proper leaf polarity. Reprinted, with permission, from the Annual Review of Plant Physiology and Plant Molecular Biology, Volume 49 (c) 1998 by Annual Reviews   www.annualreviews.org

Incipient primordia were surgically isolated from the rest of the meristem by a small incision Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

The isolated primordium lost polarity (it became entirely abaxialized) and became radially-symmetrical. P3 P2 P1 I1 I2 I3 P3 P2 P1 I1 I2 I3

A more recent experiment showed that laser ablation of only the epidermal cell layer is sufficient for the primordium to lose its adaxial polarity. Reinhardt, D., Frenz, M., Mandel, T., and Kuhlemeier, C. (2005) Development 132: 15-26. Reproduced with permission.

A signal from the meristem moves through the epidermis into the incipient primordium. The signal conveys the adaxial positional information. The nature of the signal is not known.

Controllo genetico della polarità The phantastica mutant of Antirrhinum (snapdragon) gives important clues to the basis of leaf polarity. phan Wild-type Waites, R., and Hudson, A. (1995) Development 121: 2143 – 2154. Reproduced with permission.

The phantastica mutant has radially symmetrical leaves. Wild-type leaf phan mutant leaf The phantastica mutant has radially symmetrical leaves. Waites, R., and Hudson, A. (1995) Development 121: 2143 – 2154. Reproduced with permission.

Wild-type leaf phan mutant leaf Mutant phan leaves are abaxialized, indicating that PHAN is necessary for adaxial cell fate.

Tutte le foglie radialmente simmetriche sono abaxializzate? P3 P2 P1 I1 I2 I3 Surgical isolation phan mutant leaf The phan mutant leaves resemble the surgically isolated leaf primordia –

No: I mutanti Loss of function kanadi hanno foglie radiali adaxiali A triple mutant kanadi 1,2 and 3 has radial, adaxialized leaves KANADI genes promote abaxial cell fate Eshed Y et al., Izhaki, A., Baum, S.F., Floyd, S.K., and Bowman, J.L. (2004) Development 131: 2997-3006. Reproduced with permission.

La perdita della identità adaxiale o abaxiale determina la radializzazione Wild type phan mutant kan mutant Eshed Y et al., Izhaki, A., Baum, S.F., Floyd, S.K., and Bowman, J.L. (2004) Development 131: 2997-3006. Reproduced with permission.

Fenotipo del mutante Gain-of-function phb-1d Gain-of-function phb-1d mutants have radial, adaxialized leaves. McConnell, J.R. and Barton, M.K. (1998) Development 125: 2935-2942. Reproduced with permission.

La mutazione The phb-1d mutation inluenza la distribuzione del mRNA di PHB Cross section SEM In gain-of-function phb-1d mutants, PHB is expressed everywhere, resulting in adaxialized, radially symmetric leaves. Longitudinal section Cross section In wild-type plants, PHB expression is restricted to the adaxial side of the leaves Reprinted by permission from Macmillan Publishers, Ltd: NATURE. McConnell, J.R., Emery, J., Eshed, Y., Bao, N., Bowman, J., and Barton, M.K. Nature 411: 709-713, copyright 2001.

PHB promuove l’identità adaxiale Wild-type leaf: phb-1d leaf PHB expression = Adaxial No PHB expression = Abaxial Reprinted by permission from Macmillan Publishers, Ltd: NATURE. McConnell, J.R., Emery, J., Eshed, Y., Bao, N., Bowman, J., and Barton, M.K. Nature 411: 709-713, copyright 2001.

come PHB, PHV and REV promuovono l’identità adaxiale Like PHB, REVOLUTA (REV) and PHAVOLUTA (PHV) are expressed in the adaxial domain. Reprinted from Current Biology 13, Emery, J.F., et al., Radial patterning of Arabidopsis shoots by Class III HD-ZIP and KANADI genes, 1768–177, Copyright (2003), with permission from Elsevier.

come PHB, PHV and REV promuovono l’identità adaxiale Loss of function triple phb / phv / rev mutant has radial, abaxialized leaves Reprinted from Current Biology 13, Emery, J.F., et al., Radial patterning of Arabidopsis shoots by Class III HD-ZIP and KANADI genes, 1768–177, Copyright (2003), with permission from Elsevier.

La polarità richiede la corretta espressione di PHB Too little PHB Too much PHB Borghi, L., et al.,(2007) Plant Cell 19:1795-1808.

La espressione di PHB è regolata da miRNA miRNAs are short (~21-22 nt) RNAs that, in association with ARGONAUTE (AGO) target specific mRNAs for degradation (or interfere with translation). AAAAAAA AAAAAAA AGO AAAAAAA

Controllo della espressione di PHB da parte di miRNA AAAAAAA AGO In wild-type plants, miR166 binds to the PHB mRNA and degrades it on the abaxial side of the leaf primordium. miR166 PHB mRNA AAAAAAA x PHB-1D mRNA In phb-1d plants, base changes in the PHB mRNA prevent miR166 from binding to it, allowing it to accumulate throughout the leaf primordium. Reprinted by permission from Macmillan Publishers, Ltd: NATURE. Kidner, C.A. and Martienssen, R.A. Nature 428: 81-84, copyright 2004.; McConnell, J.R., Emery, J., Eshed, Y., Bao, N., Bowman, J., and Barton, M.K. Nature 411: 709-713, copyright 2001.

I miRNA controllano la polarità della foglia Additional support for role of miRNA in leaf polarity comes from the fact that the ago1 mutant has radial leaves; AGO is needed for miR166 function. AAAAAAA AGO In ago mutants, as in phb-1D mutants, PHB mRNA accumulates throughout the leaf primordia. ago mutant phb-1D mutant Reprinted by permission from Macmillan Publishers, Ltd: NATURE. Kidner, C.A. and Martienssen, R.A. Nature 428: 81-84, copyright 2004; McConnell, J.R., Emery, J., Eshed, Y., Bao, N., Bowman, J., and Barton, M.K. Nature 411: 709-713, copyright 2001.

I miRNA contribuiscono alla determinazione della polarità della foglia These experiments demonstrate that in Arabidopsis miRNAs contribute to leaf polarity, by controlling the distribution of critical mRNAs. AAAAAAA AGO

Cosa fanno tutti questi geni? ABAXIALIZING GENES KANADI 1,2,3 encode GARP transcription factors; YABBY genes have similar function and also encode putative transcription factors ADAXIALIZING GENES PHANTASTICA encodes a MYB transcription factor; PHB/ PHV/ REV genes encode HD-ZIP III transcription factors The genes regulated by these transcription factors are not yet known.

geni YABBY (FIL, YAB2, YAB3) promuovono il differenziamento del lato abaxiale delle foglie (SONO FATTORI DI TRASCRIZIONE ZINC –FINGER) Mutanti yab3 (omozigoti) producono foglie lobate che esprimono KNOX1 e formano meristemi ectopici e mostrano conversione abaxiale/adaxiale Il fenotipo yab suggerisce che ci sia incompatibilità tra la funzione KNOX e le funzioni che specificano l’identità ABAXIALE nella foglia

Modello per l’acquisizione della polarità PHAN or PHB/PHV/REV miR166 KAN, YAB Adaxial fate Abaxial fate Meristem-derived signal

Il patterning ad/abaxiale può influenzare tratti agronomicamente importanti Wild-type rice leaf. Sclerenchymatous tissue forms on the abaxial surface and supports the leaf in an open form. Zhang, G-H. et al., (2009) Plant Cell 21:719-735

SLL1 è una proteina GARP che influenza la polarità e l’arrotolamento della foglia Wild-type sll1 mutant In sll1 mutants, the supportive sclerenchymatous tissues on the abaxial surface do not form, causing the leaf to roll inwards. Zhang, G-H. et al., (2009) Plant Cell 21:719-735

Rice plants with rolled leaves (like sll1) can have more erect leaves, reduced water loss by transpiration and higher yields. Wild-type sll1 mutant Zhang, G-H. et al., (2009) Plant Cell 21:719-735

SUMMARY Leaves are initiated from cells in the shoot apical meristem Auxin gradients are important in leaf primordium initiation and positioning Leaf identity is determined by a change in expression of transcription-factor encoding genes Leaf polarity requires an unknown signal from the meristem and the domain-specific expression of adaxial- and abaxial- specific transcription factors

GENI DI IDENTITA’ ADAXIALE/ABAXIALE

PHANTASTICA (anthirrinum)/ AS1 (arabidopsis) mutazioni loss of function phan promuovono la conversione adaxiale/abaxiale (organi a simmetria radiale) I trascritti sono localizzati nel lato adaxiale Geni di identità adaxiale I mutanti phan hanno anche difetti nel mantenimento del meristema Interdipendenza destino adaxiale / meristematico

mutazione phan

PHABULOSA, PHAVOLUTA, REVOLUTA Homeodomain leucin zipper proteins HD-ZIPIII contenenti un dominio START che lega lipidi Fattori di trascrizione specifici per il lato adaxiale

Interdipendenza destino adaxiale / meristematico PHB PHV REV Sono inizialmente espressi in maniera continua dal centro del SAM fino ai primordi fogliari; successivamente la loro espressione si restringe al lato adaxiale della foglia Mutazioni loss-of-function nei geni PHB o PHV o REV determinano conversione adaxiale/ abaxiale e incapacità di formazione o mantenimento del SAM Le funzioni PHB, PHV, REV sembrano positivamente correlate alla funzionalità dei geni KNOX Interdipendenza destino adaxiale / meristematico

Geni KANADI (KAN) espressi nella regione abaxiale La localizzazione nella regione adaxiale dei trascritti HD-ZIP III è regolata da: Geni KANADI (KAN) espressi nella regione abaxiale (mutazioni recessive KAN determinano adaxializzazione e espressione ectopica di PHAV, PHAB, REV) microRNA (mutazioni dominanti PHB e PHV inibiscono il riconoscimento e la degradazione dei trascritti genici ad opera di miRNA espressi specificamente nella regione abaxiale)

Regolazione da micro RNA dei geni HD-ZIP III miR165 miR166 Mutazioni in queste regioni danno luogo a fenotipi dominanti con foglie adaxializzate, radiali e meristemi più grandi