Plant Vacuole: Tonoplast Membrane, Atpases, Transporters As Storage Organelle

Plant Vacuole: Tonoplast membrane, ATPases,
transporters as storage organelle

• vacuole is an essential organelle for plant growth
and development.
• location for the storage of nutrients; such as sugars

and proteins; and other metabolic products.
• Understanding the mechanisms of vacuolar

trafficking and molecule transport across the
vacuolar membrane is of great importance in
understanding basic plant development and cell
biology and for crop quality improvement.
The vacuole occupies as much as 90% of most
Vacuoles mature cells.
- (coined from vacuum») – fluid-filled compartments

encompassed by a membrane called tonoplast.
10 Vacuole
Tonoplast Structure
unique to plant cells
lipid bilayer, typical of other cellular membranes.
hydrophillic (water loving) head facing out into the cellular cytosol
and the vacuolar cytosol.
 The hydrophobic (water fearing) tails face inwards.
tonoplast is semi-permeable allows the tonoplast to maintain a

balance of ions inside and outside of the central vacuole.
this maintains proper turgor pressure inside the cell.

Plant vacuoles are multifunctional
compartments
• Storage (proteins, amino acids and organic acids, ions, sugars, pigments)
• Digestion (acid hydrolases – proteases, nucleases, glycosidases, lipases)
• pH and ionic homeostasis (serve as reservoirs of protons and

metabolically important ions)
• Defense against microbial pathogens and herbivores (cell wall‐degrading

enzymes, phenolic compounds, alkaloids, etc.)
• Sequestration of toxic compounds , detoxification of xenobiotics,

leaves are unable to rid themselves of waste products or xenobiotics such as
herbicides. These are ultimately deposited in the vacuole
• Pigmentation
• recycling of cell components,

Plants use vacuoles to produce large
cells cheaply
• By filling a large volume of the cell with “inexpensive” vacuolar

contents plants are able to reduce the cost of making expanded
structures such as leaves, which are essentially solar collectors.
• space-filling function of the vacuole is essential for cell growth,

because cell enlargement is accompanied by expansion of the vacuole
rather than of the cytoplasm.
• The water taken into vacuoles generates turgor pressure which

expands the primary cell wall and creates stiff structures in
conjunction with the walls.
• part of the endomembrane system in plant cells and occupy a large
percentage of the cell volume.
• vacuolar membrane named the tonoplast separates the

cytoplasm from the vacuole lumen.
• vacuoles are very dynamic
• morphology changes in response to environmental conditions

- varies during different plant developmental stages
• Water, ions and metabolic products cannot cross the tonoplast

freely without the facilitation of tonoplast proteins.
 Controlled transport across the tonoplast -important for
plant responses to environmental conditions or intracellular
signalling
 activities of tonoplast-localized enzymes, transporters and
channels change in response to cytoplasmic conditions.
 regulate material exchange between the cytoplasm and

vacuole lumen and maintain cellular homeostasis
 Tonoplast membrane proteins can serve to facilitate and

regulate biomolecular transport across the membrane.
Types of vacuoles
Two types of vacuoles are

depicted:
large protein storage
vacuoles (V1)
and smaller
lytic/autophagic‐type
vacuoles (V2) that may be
involved in
autophagy‐associated
programmed cell death.
Biochemistry&Molecular Biology of Plants/edited by Buchmann B., Gruissem W.,
Russel L.J.
Routes towards the vacuole
• In all eukaryotes, exiting the ER is via coat protein
complex (COP)II-coated vesicles.
• The vacuole, together with the plasma membrane,

is the most distal point of the secretory pathway,
and many vacuolar proteins are transported from
the ER through intermediate compartments.
• However, the presence of alternative transport

routes from the ER towards the tonoplast, which
are independent of Golgi- and post-Golgi
trafficking.
Tonoplast
• A large number of proteins in the tonoplast support

function of multifaceted vacuoles, including active
pumps, carriers, ion channels, receptors, and
structural proteins.
• relative abundance of these proteins and their

respective activities/regulation determine the
specific function of plant vacuoles.
• the membrane delimiting plant vacuoles, regulates

ion, water and nutrient movement between the
cytosol and the vacuolar lumen through the activity
of its membrane proteins
The tonoplast is a functionally, highly organized membrane,
with proton pumps and several H+/X antiporters, which use
a pH gradient generated by proton pumps, cooperatively
working in the same membrane.
Thus, there is a proton circuit on the tonoplast.
Various ion channels also utilize the membrane potential

generated by proton pumps
cooperative work of these different transport systems

regulates the vacuole lumenal pH, amount of stored
substances, and volume of the vacuole.
The three most abundant proteins of the tonoplast are
 vacuolar H+-ATPase (V-ATPase), composed of several subunits, is

the largest complex in the tonoplast.
 H+-pyrophosphatase (V-PPase), an alternative proton pump, is also

a major component of the tonoplast in most plant tissues
1. These pumps acidify vacuoles and generate an electrochemical gradient

in the plant cell.
2. This proton electromotive force allows the secondary active transport of

inorganic ions, sugars, and organic acids
 water channels (aquaporins). tonoplast intrinsic protein.

vacuolar H+-ATPase (V-ATPase)
 V-ATPase -largest complex in tonoplast
 molecular size 750 kDa.

 composed of two functional sectors
a peripheral sector (V1)- catalytic sites for ATP
hydrolysis
transmembrane sector (VO) a channel for protons.
 catalytic portion (10–12 nm in diameter) of V1 consists

of three copies of A- and B-subunits.
 other subunits (C–H) in V1 form a central stalk
(6–7 nm height) linking the V1 and VO sectors.
 VO sector contains the a- and c-subunits (16 kDa)

TONOPLAST TRANSPORT MACHINERY Hypothetical model for the
tertiary structure of plant V-ATPase. The V1 catalytic domain consists of
subunits A and B in an A3B3 arrangement. The VO membrane sector is
composed of a- and c-subunits in the ac6 complex.
H+-pyrophosphatase (V-PPase),
• essential for maintaining the acidity of the large central vacuole
METAL ION TRANSPORTERS Vacuolar Ca2+-ATPase

 vacuole serves as a primary pool of free calcium ions in plant cells -
major source of Ca2+ for intracellular calcium signalling.
 The Ca2+- ATPase and Ca2+/H+ antiporter perform active Ca2+

transport into the vacuole
A transmembrane domain containing a Ca2+-binding site

and three cytoplasmic domains (A, N and P) of the Ca2+-
ATPase were recognized in the molecular architecture. A
phosphorylation site is located in domain P, and the
adenosine moiety of ATP is bound to domain N. P
Vacuolar Ca2+/H+ Antiporter
1.consist of a single polypeptide of 399–444 amino acid

residues.
 The Ca2+/H+ antiporter together with the Ca2+-ATPase

plays a key role in vacuolar Ca2+ accumulation.
 antiporter driven by transmembrane pH gradient
generated by the V-ATPase and V-PPase.
Vacuolar Na+/H+ Antiporter 538 amino acid residues
export Na+ from the cytosol both to the extracellular space

and to the vacuole driven by proton gradients across the
plasma membrane and tonoplast
Zinc Transporter
Zinc is an essential plant micronutrient and functions as
an essential cofactor for many enzymes: alcohol
dehydrogenase, carbonic anhydrase, carboxypeptidase,
DNA polymerase, RNA polymerase, and transcription
factors with zinc
fingers
AQUAPORIN
 Aquaporins facilitate water transport across biomembranes in
an osmotic pressure dependent manner.
 belong to a ubiquitous family of membrane intrinsic proteins

homotetramer of subunits of 23–30 kDa
 A high water permeability for the tonoplast, facilitated
 important in protecting the cell against plasmolysis.
 Enables vacuoles to transport water quickly and in large

quantities to the cytosol, even with a small osmotic gradient
across the tonoplast.
AQUAPORIN
V-ATPase, V-PPase, and TIP form a 750-kDa complex, a dimer,
and a tetramer in the tonoplast,
Transport processes across the
tonoplast
•The regulation of solute passage across

the tonoplast can be achieved by
modified expression of genes encoding
tonoplast proteins. However, also post-
translational modifications of tonoplast
proteins represent a fundamental
principle in vacular transport regulation
and adaptation.
H. Ekkehard, O. Trentmann, Regulation of transport processes across the tonoplast,

September 2014
Schematic drawing illustrating the current knowledge on

how tonoplast monosaccharide transporters (TMTs) are
regulated at the post-translational level.
Autophagy
• is a degradation pathway that recycles cell materials upon stress
conditions or during specific developmental processes (in the
lysosome for mammals or in the vacuole for yeast and plants).
Thank you for attention!

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Plant Vacuole: Tonoplast membrane, ATPases,

transporters as storage organelle

• location for the storage of nutrients; such as sugars

• Understanding the mechanisms of vacuolar

- (coined from vacuum») – fluid-filled compartments

unique to plant cells

lipid bilayer, typical of other cellular membranes.

 The hydrophobic (water fearing) tails face inwards.

tonoplast is semi-permeable allows the tonoplast to maintain a

this maintains proper turgor pressure inside the cell.

• Digestion (acid hydrolases – proteases, nucleases, glycosidases, lipases)

• pH and ionic homeostasis (serve as reservoirs of protons and

• Defense against microbial pathogens and herbivores (cell wall‐degrading

• Sequestration of toxic compounds , detoxification of xenobiotics,

• recycling of cell components,

• By filling a large volume of the cell with “inexpensive” vacuolar

• space-filling function of the vacuole is essential for cell growth,

• The water taken into vacuoles generates turgor pressure which

• vacuolar membrane named the tonoplast separates the

• vacuoles are very dynamic

• morphology changes in response to environmental conditions

• Water, ions and metabolic products cannot cross the tonoplast

 regulate material exchange between the cytoplasm and

 Tonoplast membrane proteins can serve to facilitate and

Two types of vacuoles are

• The vacuole, together with the plasma membrane,

• However, the presence of alternative transport

• A large number of proteins in the tonoplast support

• relative abundance of these proteins and their

• the membrane delimiting plant vacuoles, regulates

Thus, there is a proton circuit on the tonoplast.

Various ion channels also utilize the membrane potential

cooperative work of these different transport systems

 vacuolar H+-ATPase (V-ATPase), composed of several subunits, is

 H+-pyrophosphatase (V-PPase), an alternative proton pump, is also

1. These pumps acidify vacuoles and generate an electrochemical gradient

2. This proton electromotive force allows the secondary active transport of

 water channels (aquaporins). tonoplast intrinsic protein.

 molecular size 750 kDa.

 catalytic portion (10–12 nm in diameter) of V1 consists

 VO sector contains the a- and c-subunits (16 kDa)

METAL ION TRANSPORTERS Vacuolar Ca2+-ATPase

 The Ca2+- ATPase and Ca2+/H+ antiporter perform active Ca2+

A transmembrane domain containing a Ca2+-binding site

1.consist of a single polypeptide of 399–444 amino acid

 The Ca2+/H+ antiporter together with the Ca2+-ATPase

export Na+ from the cytosol both to the extracellular space

 belong to a ubiquitous family of membrane intrinsic proteins

 A high water permeability for the tonoplast, facilitated

 important in protecting the cell against plasmolysis.

 Enables vacuoles to transport water quickly and in large

•The regulation of solute passage across

H. Ekkehard, O. Trentmann, Regulation of transport processes across the tonoplast,

Schematic drawing illustrating the current knowledge on

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