Plant Growth Control : Physical Methods

Kinkajou : Let’s talk about methods of controlling plant growth.

Erasmus :Plant growth can be controlled by physical methods, pruning, environmental factors and chemicals such as plant growth hormones.


One physical method is staking. This is the practice of using vertical stakes to support trees whose growth habit or rapid growth does not enable them to support themselves. Rapid growth is desirable in timber species trees as this produces straight wood suitable for building applications. Done inappropriately, rubbing against rope tenders or the stake itself stimulates cell growth resulting in deformity of the trunk and thickening around the straps.

Staking Tomatoes- Growth Control

  
 Pruning

 Pruning is a method whereby a tree can be shaped by removal of the shoot apex (dominant) or lateral stems. Removal of the shoot apex (typically the tallest part of the plant) eliminates the dominance of the apex over stem buds below. This allows the buds to form new branches, effectively thickening the canopy of the tree or shrub.
If an entire branch of the lateral lower part of the tree or shrub is removed, this is known as “thinning”. If only a third or a half of branch is removed, the process is called “heading back”.

If the entire trunk of the tree or shrub is cut, this is known as a “heading (or topping) cut”. Often new shoots form from just below the cut line but these are weakly attached to the trunk, increasing the probability of new branches breaking off is the tree grows. Appropriate pruning allows farmers to control and direct future plant growth, and to train them into specific shapes. For example many fruit trees have the apex truncated and pruned so that the top of the tree is often slightly concave. This allows easier harvesting of fruit crops.

Pruning Plants Cuts

Goo :So controlling plant growth is not all about genes and chemicals.

Erasmus :Right on.   But plant hormones are well recognised and have been able to be used commercially. In horticulture hormones and plant growth regulators are commonly used to control plant growth habit. These chemicals are incredibly potent, so often are only relevant in commercial not residential applications.

Pruning Growth Control

 

 

Plant Growth Control : Plant Hormones

Kinkajou : Tell us about plant hormones.

Erasmus :
Auxin Hormones are produced in the terminal buds of plants. Typically they suppress the growth of side buds and stimulate root growth. They also influence cell division and differentiation as well as cell elongation. The effect is to make the tree taller by promoting apical dominance, giving young tree is a more upright form. Auxin levels reduce with maturity resulting in trees developing a more rounded Crown and stopping the growth of the tree in height.

Pruning a plant plan

Gibberellin hormones affect the rate of cell division, flowering and seed or Bud dormancy. They also affect the size of leaves and fruit. They are commonly used to induce plant growth at low temperatures where the plant would typically stay within a more dominant state.

Exposure time and exposure concentration are all variables which can be manipulated to control cell growth. As plants metabolise at different rates in the morning versus the evening, on cloudy days, and in different humidity conditions, these can also affect the effect of these hormones when use commercially. Some of these agents and bond to organic chemicals, so again growth medium can result in changes in the degree of action.

Gibberellin Hormone- Plant Growth Effects

Erasmus :   Other chemicals affecting plant growth include:

• Brassinosteroids
• Xylogens
• Cytokinins promote cell division, and influence cell differentiation and aging of leaves.
• Abscisic acid is considered the “stress” hormone.  It inhibits the effects of other hormones to reduce growth during times of plant stress.
  
  

Auxins and Gibberellins are the reasons why pruning adjusts plant growth. Pruning the top of a tree removes the Auxin, changing the growth habit as well as slowing root generation. “Heading cuts” release Auxins, allowing side shoots to develop and the branch or trunk to become bushy. Heading cuts reduce apical dominance and the branches become denser and lateral buds begin to grow. Auxins also affect the direction of plant stem growth (called tropism). A tree balances canopy growth against root growth by controlling the levels of Auxins and Gibberellins.

The obverse process to heading cuts is the making of thinning cuts. A thinning cut removes a branch at a branch union (aka crotch). It opens the plant for better light penetration. By redirecting the nutrients/sugars to terminal shoots, an open growth habit is promoted.

Geotropism Plants

Plant cells contain structures called amyloplasts that sediment under gravity. This is sensed within the cell causing Auxins to accumulate in the lower side of a horizontal stem, resulting in some cells to enlarging faster, turning the stem upright. (This is called geotropism). Another form of tropism allows the plant to turn towards a light source. Auxins accumulate on the shady side of the tree. They are reduced on the sunny side. This causes plant cell elongation turning the plant towards the sun. It is the position of the Auxins within the plant that is important in determining growth pattern.

Plants can be trained to grow in many directions if you understand them.

Tropism
Auxins affect the direction of plant growth. This process is called tropism, In Geotropism, gravity is sensed by plant cells. Auxins accumulate in the lower side of a horizontal stem. Cells here consequently enlarge faster, in effect turning the stem upright. In Phototropism, Auxins increase on the shaded side of the plant. This causes cell elongation on this side, effectively turning the plant towards the sun.

Phototropism

Kinkajou : In commercial agriculture, do we use hormonal methods to control plant growth?

Erasmus :Adjusting plant size is important in the production of crops in greenhouses. There are a number of different methods.
By selecting short cultivars (dwarf varieties) plant height can be controlled. Genetics programs the hormone distribution in the plant.

Plant height can also be reduced by increasing light intensity, minimising light at the red and the spectrum. Reducing temperature variation between day and night can also be effective. The effectiveness of the strategies is plant specific.

Plant growth regulators are also used. Again their actions are plant specific due to variable modes of action. Many plant growth regulators (PGRs) act by blocking Gibberellin (GA) production.

Phototropism explanation


Erasmus :What follows is a list of PGRs and a summary of their actions.
,

• Daminozide
  Daminozide renders a key enzyme for GA production useless, thus reducing GA levels.
  
• Chlormequat Chloride
  Chlormequat chloride (inhibits GA production early in the process).
  
• Ancymidol, Flurprimidol, Paclobutrazol, and Uniconazole
  Ancymidol, flurprimidol, paclobutrazol, and uniconazole have similar helical structures. They all inhibit GA production at similar sites in the GA production process.
  
• Ethephon
  Unlike other PGRs, ethephon does not inhibit GA production. Plants take up ethephon through the leaves where it is converted to ethylene in plant cells. The increased ethylene causes cells to limit elongation and increase in width instead.
  

Ethephon’s mode of action can offer benefits other than height suppression. The release of ethylene reduces apical dominance, which can increase axillary branching. However, if the application is made close to flowering, the ethylene can result in flower abortion and delayed flowering.

Plant Hormone Growth effects

 

Plant Structure and Plant Complexity

Kinkajou : Do plant cells undergo specialisation (differentiation), much like the cells in different organs in animals?

Erasmus : What is interesting about plant cells is that in some plants cells can undergo a program of the de-differentiation and re-differentiation. This can allow a fragment of plant to regenerate the entire plant. In short, each plant cell retains the genetic material and the capacity to regenerate the entire plant. This is not true for differentiated animal cells.

Sprouting Stump Regenerating

Kinkajou : So there is a difference between plant and animal cells, but perhaps the problem is just that we are more familiar with animal cell growth regulation. There may be restrictions in plant cell differentiation that we simply are not aware of, especially if the process is species specific.






Erasmus :Let’s keep going to describe the cellular structure of plants, something most of us are not very familiar with. The parts of a plant include:
Plant Stems / Roots are composed of concentrically organized cell layers.
From in­side to outside these are

• vasculature  > stele
• pericycle>stele 
  The vasculature and peri-cycle together constitute the stele

       

• endodermis >
  Parts of this cell layer grow to form the Xylem and the phloem tissues
• cortex  > Trichomes and stomata
• epidermis >epidermis

. Plant Stem Structure Macro

 

 

Three main cell types exist in plant tissues. These are

• stomata,
• Trichomes and
• Epidermal pavement cells.
  
  
• Stomata are specialised cells. They control the gas exchange through pores formed by guard cell pairs.
• Trichomes can be very complex in some plants. They may be unbranched or stellate, unicellular or multicellular, and often plant hairs differentiate into specialized secreting cells. They can form hair like structures allowing the plant to protect itself from dehydration, excess light intensity and perhaps even against insects.  
  .
• Specialised cells in plants include the epidermis. This is the outermost layer of plant. Cells typically have a thick apical wall on the exterior that secretes an impermeable external wax layer. In branches and stems (aerial tissues) it is important for gas exchange. In root structures is important for nutrient uptake.
  

The shoot apical meristem (SAM) is the source of cells for all aerial organs produced after germination.  

 

Plant Stem Substructure2

Compared with the SAM (Shoot Apical Meristem), the root meristem (RM) has fewer cells and a simpler structure.  Cells in each layer have their origin in the “initial cells” located at the RM (root meristem), which re­peat a highly stereotyped sequence of divisions. One of the two daughter cells remains as an initial cell, whereas the other enters an appropriate differentiation pathway. Root hairs develop from trichoblasts.
Two plant hormones Auxin and ethylene are responsible for the control of much of the cell differentiation.


Kinkajou : So describe the process of plant cell growth / differentiation.

Erasmus :When a plant germinates, early root meristem (RM) cells are activated in genetically determined cell division sequences. These give each plant the typical root distribution pattern of its species. Research suggests that root distribution and formation requires a “top-down “flow of positional information. It appears that Auxins may have a role in root patterning.

Other forms of molecular signalling are important in the internal plant environment including the deeper layers of the plant stem. These can involve special signalling proteins, like receptor –ligand interactions and movement of transcription factors across cell layers. Recent biotechnology advances have resulted in key regulatory genes being identified based on visible phenotypic alterations. Each cell appears to produce signalling factors (of proteins) that affect the growth of other cell types adjacent in an interdependent fashion. Laser ablation study to different cell layers and growing root has shown substantial changes in the pattern of cell growth in the newly forming root.

Kinkajou : We’ve talked about the aerial (above ground) and the root (below ground) structures. What about the plant vascular system?

Erasmus :There are two main cells types existing in the plant vascular system. These are Xylem cells and Phloem cells,
Xylem cells are highly specialized cells of the vascular system required for the regulated transport of water and nutrients. They can be induced to form from parenchymal cells by wound stress or plant hormones.
Initially, parenchymal cells dedifferentiate into a xylem precursor cells without undergoing cell division.

• During the initial stage, parenchymal cells dedifferentiate without undergoing cell division, differentiating into a xylem precursor cell.
  
• The second stage is defined a general increase of transcriptional and physiological activities including an increase of the levels of the endoplasmic reticulum (ER), vesicle formation, mitochondria and tubulin.
  
• In the third final stage, xylem cells form secondary cell walls and enter programmed cell death. The spatially ordered deposition of secondary cell walls depends on the coordinated organization of actin and microtubules which are thought to guide the movement of cellulose-synthesizing complexes in the plasma membrane. The secondary cell walls undergo rigidification in response to specific signalling proteins and enzymes promoting lignin biosynthesis. As the cells die, the nucleus and other organelles within the cell structure disappear, resulting in a mature transport element.

 

Erasmus :Phloem Cells form in the innermost layer of the plant stem. They are responsible for transporting organic nutrients and signalling factors particularly simple carbohydrates (sugars) to parts of the plant where needed. Phlegm cells exist as series of connected sieve cells that form a syncytium.
Many cellular elements are controlled by signalling factors. These include actin microfilaments and, microtubule-based microfilaments. So expansion is thought to be regulated by a balance between localised cell wall loosening and internal cellular hydrostatic pressure (turgor), which provides the drive for expansion.

Kinkajou : So plants are starting to look quite complicated.

Erasmus :Well you would have to expect that. Plants have been evolving for billions of years. Apparent simplicity in plant function and growth is a result of their narrow functional niche. They grow, photosynthesize and have limited motor functions. There is likely just as much genetic complexity underlying plant cells as there is in animal cells. Animal cells just look a bit different from animal to animal and tissue to tissue. So the complexity is just a bit more superficially evident.

 

Animal Cell Growth Control

Kinkajou : So let’s talk about Animal Cell Growth now.

Erasmus :Most cells in the human body exist in phase G0. Fully differentiated cells such as neurons and skeletal muscle have completely non-functional cell replication systems. Many other cells retain some capacity to undergo replication, even though differentiated. (E.g. liver cells)

One of the key regulatory factors in growth, is basal CDK (cyclin -dependent kinase) inhibition. CDK is important in regulating transcription, mRNA processing and in some cell differentiation. In the absence of mitogenic signal to proliferate, CDK inhibition is maintained and the cell remains in a non-growing state G0.

 

 

Animal Cell Growth Control: Initiation

Erasmus :Steps Animal Cell Growth: Initiation

• mitogen signalling in G1 by attaching to receptors on the cell surface
• activation of GTPase Ras
• activation of the MAP kinase cascade (Cyclin E in combination with a kinase subunit (Cdk2), is important in mammalian cells)
• Increasing levels of the gene regulatory protein Myc which promotes cell cycle entry by a number of mechanisms.
• Increase in as transcription of genes that encode for G1 phase cyclins (
• stimulating transcription of the E2F gene leading to activation of this gene regulatory protein
• Phosphorylation of the inhibitory protein Rb (retinoblastoma protein). (The cell cannot proliferate until it gets a signal from the ‘starter’ enzyme, cyclin D-dependent kinase, which phosphorylates RB and lowers RB's activity.)

Erasmus :These mechanisms are thought to operate in a double -negative feedback loop, (neg plus neg equals pos), creating a bi-stable switch. This picture of the cell-death decision as a one-way bistable switch is still speculative; there is no firm experimental evidence—either pro or con—that the signalling pathway for apoptosis has two stable steady states (‘living’ and ‘dying’). However, it appears that the pathway has a stable ‘off’ state, because living cells do not spontaneously commit suicide in response to inevitable small fluctuations of the regulatory proteins.

Furthermore, the commitment to apoptosis (flipping to the ‘on’ state) appears to be a point of no return, because cells do not revert to the ‘living’ state if the death signals are later withdrawn.

Kinkajou : So where does this all lead?

Erasmus :The final result of these changes is the activation of genes required for entry into mitosis. Many of the genes involved in replication have been identified as cancer promoting genes or oncogenes. Single amino acid Ras mutations can causes the protein to have an increased biological effect (overactivity). Mutations causing an overexpression of Myc also promote excessive cell growth and proliferation. However, when Ras or Myc is experimentally hyperactivated, cells do not undergo proliferation. They undergo cell cycle arrest or apoptosis. It therefore appears that normal cells are able to detect abnormal mitogenic stimulation and to block the effects.

 

 

Hayflick Limit for Animal Cell Growth Control

The Hayflick limit of fibroblasts in culture represents an intracellular mechanism which slows down and eventually halts fibroblasts proliferation. Normal human Fibroblasts will only go through about 25 to 50 cycles of replication when cultured in a standard mitogenic media. They then undergo a form of senescence and will not replicate.



Kinkajou : So how do we explain the Hayflick limit phenomenon?

Erasmus :Explanations for these phenomena include:

• A progressive increase in CKI (cyclin kinase inhibitor) proteins
• Telomere depletion (Telomere replication occurs independently of the replication of the remainder of the cellular DNA genome. The DNA sequences at the end of chromosomes i.e. the telomeres are synthesised by the enzyme telomerase which also forms a protein cap structure on the chromosomal end).

 

Animal Cell Growth Control: Growth

Kinkajou : So the next step after initiation in animal cell systems is growth. Tell us more.

Erasmus :Steps of Animal Cell Growth: Growth

• PI 3-kinase is one of the main intracellular signalling pathways activated by growth factor receptors
• Activation of PI 3-kinase, which promotes protein synthesis,
• Myc stimulates cell metabolism and cell growth by increasing transcription of genes associated with replication and protein synthesis.
• PDGF can act both as a growth factor as well as on mitogen.
• The signalling protein Ras can be activated by growth factors as well as mitogens.

It is believed that the dual activation of mitogens and growth factors helps to ensure that cells maintain their size as they proliferate.

Category: Oncogene Promoters	Examples
Growth factors, or mitogens	c-Sis
Receptor tyrosine kinases	epidermal growth factor receptor(EGFR),  platelet-derived growth factor receptor (PDGFR), vascular endothelial growth factor receptor (VEGFR),  HER2/neu
Cytoplasmic tyrosine kinases	Src-family, Syk-ZAP-70 family,  BTK family of tyrosine kinases, the Abl gene in CML -Philadelphia chromosome
Cytoplasmic Serine/threonine kinases  and their regulatory subunits	Raf kinase, and cyclin-dependent kinases  (through overexpression).
Regulatory GTPases	Ras protein
Transcription factors	myc gene

 

(The situation is complex because often these proteins, enzymes have different names).

Animal Cell Growth Control: Survival Factors

Kinkajou : Are there any other factors we believe are important in controlling cell growth?

Erasmus : Animal cells require signals via extracellular “survival factors” to survive or grow. Such factors include:

• Contact with a surface: Cells in culture (which are not attached to a solid surface), Almost never divide. This phenomenon is called Anchorage dependence. It is thought that the connection of extracellular matrix molecules such as laminin or fibronectin to a surface, leads to the local activation of protein kinases such as focal adhesion kinase (FA K) leading to activation of intracellular pathways promoting cell survival, growth and replication.

• Contact with other cells: when cells in culture dish form a monolayer adherent of the bottom of the dish, they normally stop proliferating. This phenomena is known as “density -dependent inhibition” of cell division. It is believed to be due to the depletion in the culture medium of extracellular mitogens. Culture Cells maintained with fresh medium flowing over their surface, grow in a different fashion to a greater population density than simple monolayer cells.
  
• Inhibitory signal proteins such as TGF-β which inhibits the proliferation of several cell types, either by blocking cell-cycle progression in G1 or by causing apoptosis. One protein in this family of proteins namely “myostatin” have been determined to act to inhibit myoblasts replication and growth in skeletal muscle cells in cattle. Some cattle breeds grown for large muscle mass have been discovered to have mutations of this gene.
  
• Survival factors: there may be other factors yet unknown which act to control cell growth and proliferation.
  
  
  

Kinkajou : So mitogens and growth factors are involved in animal cell growth control.

Erasmus :Yes!  In many multicellular organisms, mitogens and growth factors act to control total body mass which depends not only on cell numbers but also cell size. Much of the process of foetal development of attributes such as limbs and organs of specific size and shape depends on complex control system interactions with local concentrations of extracellular signal proteins stimulating or inhibiting cell growth, replication or survival. It will be some time before we understand how these genes and other undiscovered genes regulate growth and differentiation to generate a complex organism.

 

Goo :Understanding cell growth is obviously an important factor for the future. It will help us understand the immune system. It will help us control plant growth and food production. Perhaps one day in may even help us to colonise the oceans. Cellular cloning is an important tool in this process.