Editing Protoplast

{{Short description|Cell stripped of cell-wall}}
{{for|mythological progenitors of humanity|Protoplast (religion)}}
{{cs1 config |name-list-style=vanc |display-authors=6}}
[[File:Protoplasts Petunia sp.jpg|thumb|right|Protoplasts of cells from a petunia's leaf]]
[[File:Physcomitrella protoplasts.jpg|thumb|Protoplasts of the moss ''[[Physcomitrella patens]]'']]
'''Protoplast''' ({{etymology|grc|''{{wikt-lang|grc|πρωτόπλαστος}}'' ({{grc-transl|πρωτόπλαστος}})|first-formed}}), is a [[biology|biological]] term coined by [[Johannes von Hanstein|Hanstein]] in 1880 to refer to the entire cell, excluding the cell wall.<ref>{{cite book | vauthors = von Hanstein JL | title = Das Protoplasma als Träger der pflanzlichen und thierischen Lebensverrichtungen für Laien und Fachgenossen. | publisher = Selbstverlag | location = Heidelberg | date = 1880 | url = https://archive.org/details/DasProtoplasma }}</ref><ref>{{cite book | vauthors = Sharp LW | title = An introduction to cytology. | location = New York | publisher = McGraw-Hill book Company, Incorporated | date = 1921 | url = https://archive.org/details/introductiontocy032473mbp | page = 24 }}</ref>  Protoplasts can be generated by stripping the [[cell wall]] from [[plant]],<ref name="Davey-2005" /> [[bacterium|bacterial]],<ref name="Lamb, AJ-2016" /><ref name="www.encyclopedia.com-2016" /> or [[fungus|fungal]] cells<ref name="www.encyclopedia.com-2016" /><ref name="Hoondal, GS-2006" /> by mechanical, chemical or enzymatic means.

Protoplasts differ from [[spheroplasts]] in that their cell wall has been completely removed.<ref name="Lamb, AJ-2016">{{cite journal | vauthors = Cushnie TP, O'Driscoll NH, Lamb AJ | title = Morphological and ultrastructural changes in bacterial cells as an indicator of antibacterial mechanism of action | journal = Cellular and Molecular Life Sciences | volume = 73 | issue = 23 | pages = 4471–4492 | date = December 2016 | pmid = 27392605 | pmc = 11108400 | doi = 10.1007/s00018-016-2302-2 | hdl-access = free | s2cid = 2065821 | hdl = 10059/2129 }}</ref><ref name="www.encyclopedia.com-2016">{{cite web |url=https://www.encyclopedia.com/science/encyclopedias-almanacs-transcripts-and-maps/protoplasts-and-spheroplasts |title=Protoplasts and spheroplasts |publisher=Encyclopedia.com |date=2016 |website=www.encyclopedia.com |access-date=July 21, 2019}}</ref> Spheroplasts retain part of their cell wall.<ref name="www.merriam-webster.com-2019">{{cite web |url=https://www.merriam-webster.com/dictionary/spheroplast |title=Definition of spheroplast |publisher=Merriam-Webster |date=2019 |website=www.merriam-webster.com |access-date=July 21, 2019}}</ref> In the case of [[Gram-negative bacteria]]l spheroplasts, for example, the [[peptidoglycan]] component of the cell wall has been removed but the [[bacterial outer membrane|outer membrane]] component has not.<ref name="Lamb, AJ-2016" /><ref name="www.encyclopedia.com-2016" />

==Enzymes for the preparation of protoplasts==
Cell walls are made of a variety of [[polysaccharide]]s.  Protoplasts can be made by degrading cell walls with a mixture of the appropriate polysaccharide-degrading [[enzyme]]s:
{| class="wikitable"
!Type of cell
!Enzyme
|-
| [[Plant cell]]s
|[[Cellulase]], [[pectinase]], [[xylanase]]<ref name="Davey-2005" />
|-
| [[Gram-positive]] bacteria 
| [[Lysozyme]], ''N,O''-diacetylmuramidase, [[lysostaphin]]<ref name="Lamb, AJ-2016" />
|-
| [[Fungus|Fungal]] cells
| [[Chitinase]]<ref name="Hoondal, GS-2006">{{cite journal | vauthors = Dahiya N, Tewari R, Hoondal GS | title = Biotechnological aspects of chitinolytic enzymes: a review | journal = Applied Microbiology and Biotechnology | volume = 71 | issue = 6 | pages = 773–782 | date = August 2006 | pmid = 16249876 | doi = 10.1007/s00253-005-0183-7 | s2cid = 852042 }}</ref>

|}
During and subsequent to digestion of the cell wall, the protoplast becomes very sensitive to [[osmosis|osmotic]] stress.  This means cell wall digestion and protoplast storage must be done in an [[Tonicity#Isotonicity|isotonic solution]] to prevent rupture of the [[plasma membrane]].{{Citation needed|date=August 2023}}

==Uses for protoplasts==
[[File:Protoplast fusion.jpg|thumb|right|Fused protoplast (on left), containing both chloroplasts (from a leaf cell) as well as a  coloured vacuole (from a petal).]] 
Protoplasts can be used to study [[biological membrane|membrane]] biology, including the uptake of [[macromolecule]]s and  [[virus]]es . These are also used in [[somaclonal variation]].

Protoplasts are widely used for [[DNA]] [[Transformation (genetics)|transformation]] (for making [[genetically modified organism]]s), since the cell wall would otherwise block the passage of DNA into the cell.<ref name="Davey-2005">{{cite journal | vauthors = Davey MR, Anthony P, Power JB, Lowe KC | title = Plant protoplasts: status and biotechnological perspectives | journal = Biotechnology Advances | volume = 23 | issue = 2 | pages = 131–171 | date = March 2005 | pmid = 15694124 | doi = 10.1016/j.biotechadv.2004.09.008 }}</ref> In the case of plant cells, protoplasts may be regenerated into whole plants first by growing into a group of plant cells that develops into a [[Callus (cell biology)|callus]] and then by regeneration of shoots ([[caulogenesis]]) from the callus using [[plant tissue culture]] methods.<ref>{{cite journal | vauthors = Thorpe TA | title = History of plant tissue culture | journal = Molecular Biotechnology | volume = 37 | issue = 2 | pages = 169–180 | date = October 2007 | pmid = 17914178 | doi = 10.1007/s12033-007-0031-3 | s2cid = 25641573 }}</ref> Growth of protoplasts into callus and regeneration of shoots requires the proper balance of plant growth regulators in the tissue culture medium that must be customized for each species of plant.<ref>{{cite journal | vauthors = Sandgrind S, Li X, Ivarson E, Ahlman A, Zhu LH | title = Establishment of an Efficient Protoplast Regeneration and Transfection Protocol for Field Cress (<i>Lepidium campestre</i>) | journal = Frontiers in Genome Editing | volume = 3 | pages = 757540 | date = 16 November 2021 | pmid = 34870274 | pmc = 8635052 | doi = 10.3389/fgeed.2021.757540 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Li X, Sandgrind S, Moss O, Guan R, Ivarson E, Wang ES, Kanagarajan S, Zhu LH | title = Efficient Protoplast Regeneration Protocol and CRISPR/Cas9-Mediated Editing of Glucosinolate Transporter (<i>GTR</i>) Genes in Rapeseed (<i>Brassica napus</i> L.) | journal = Frontiers in Plant Science | volume = 12 | pages = 680859 | date = 2021-07-07 | pmid = 34305978 | pmc = 8294089 | doi = 10.3389/fpls.2021.680859 | doi-access = free | bibcode = 2021FrPS...1280859L }}</ref> Unlike protoplasts from [[vascular plant]]s, protoplasts from [[moss]]es, such as ''[[Physcomitrella patens]]'', do not need [[phytohormone]]s for regeneration, nor do they form a callus during [[Regeneration (biology)|regeneration]]. Instead, they regenerate directly into the filamentous [[protonema]], mimicking a germinating moss spore.<ref>Bhatla SC, Kiessling J, [[Ralf Reski|Reski R]] (2002): [https://archive.today/20130203001100/http://www.springerlink.com/content/m747f7xfu9dxyprw/?p=2747894a7a9849a09e60b0e49f51afdc&pi=3 Observation of polarity induction by cytochemical localization of phenylalkylamine-binding receptors in regenerating protoplasts of the moss ''Physcomitrella patens'']. ''Protoplasma'' 219, 99–105.</ref>

Protoplasts may also be used for [[plant breeding]], using a technique called [[protoplast fusion]].  Protoplasts from different species are induced to fuse by using an [[electric field]] or a solution of [[polyethylene glycol]].<ref>{{cite journal | vauthors = Hain R, Stabel P, Czernilofsky AP, Steinbiß HH, Herrera-Estrella L, Schell J | title = Uptake, integration, expression and genetic transmission of a selectable chimaeric gene by plant protoplasts. | journal = Molecular and General Genetics MGG | date = May 1985 | volume = 199 | issue = 2 | pages = 161–168 | doi = 10.1007/BF00330254 }}</ref> This technique may be used to generate [[somatic hybrid]]s in tissue culture.{{Citation needed|date=August 2023}}

Additionally, protoplasts of plants expressing fluorescent proteins in certain cells may be used for Fluorescence Activated Cell Sorting (FACS), where only cells fluorescing a selected wavelength are retained. Among other things, this technique is used to isolate specific cell types (e.g., [[guard cells]] from leaves, [[pericycle]] cells from roots) for further investigations, such as transcriptomics.{{Citation needed|date=August 2023}}

== See also ==
*[[Bacterial morphological plasticity]]
*[[L-form bacteria]]
*[[Spheroplast]]s

== References ==
{{Reflist}}

{{Authority control}}

[[Category:Cell biology]]
[[Category:Membrane biology]]
[[Category:Molecular biology]]
[[Category:Plant physiology]]
[[Category:Plant reproduction]]