Pal Maliga

Last updated
Pal Maliga
Pal Maliga 2020.jpg
Pal Maliga, Piscataway, NJ, in 2020
Born(1946-02-23)23 February 1946
Scientific career
Fields Agrobacterium engineering, Expression of recombinant proteins in Chloroplast, CRISPR/Cas for organellar genome engineering
Institutions

Pal Maliga is a plant molecular biologist. He is Distinguished Professor of Plant Biology and Laboratory Director at the Waksman Institute of Microbiology, Rutgers University. He is known for developing the technology of chloroplast genome engineering in land plants and its applications in basic science and biotechnology.

Contents

Research

Chloroplast genome engineering

The Maliga group in Szeged isolated chloroplast-encoded antibiotic-resistance [1] [2] [3] [4] and herbicide-resistant mutants [5] in cultured tobacco cells and have shown that chloroplast-encoded antibiotic-resistance gives a selective advantage to chloroplasts in cultured cells. [6] The ability to selectively enrich resistant chloroplasts was the foundation for obtaining chloroplast genome-engineered (transplastomic) tobacco plants. [7] Extensive recombination of chloroplast genomes after chloroplast fusion confirmed homologous recombination in chloroplasts, [8] [9] providing a template for the design of chloroplast transformation vectors. The Maliga laboratory achieved tobacco (Nicotiana tabacum) chloroplast genome transformation in 1990 by selection for spectinomycin resistance encoded in the 16S rRNA, a process that was made efficient by selection for chimeric antibiotic resistance genes. [10] [11] [12] The significance of chloroplast genome engineering as a tool to improve photosynthetic efficiency was recognized early on. [13] In arabidopsis (Arabidopsis thaliana) efficient chloroplast transformation required knocking out a nuclear gene. [14] The toolkit for chloroplast genome engineering was completed by post-transformation excision of marker genes using phage site-specific recombinases. [15]

Agrobacterium transformation

The Maliga team constructed the pPZP Agrobacterium binary vector family, [16] that served as the backbone for the CAMBIA and GATEWAY Agrobacterium vectors. Currently they are engaged in reengineering Agrobacterium for DNA delivery to chloroplasts, [17] so that chloroplast transformation can be achieved by the floral dip protocol.

Chloroplast transcription

Chloroplast reverse genetics revealed the distinct role of two plastid RNA polymerases. [18] [19] The Maliga lab characterised plastid promoters in vivo and in vitro, and identified proteins that are parts of the plastid PEP transcription complex. [20]

Expression of recombinant proteins in chloroplasts

One of the first biotechnological applications of chloroplast engineering was expression of Bacillus thuringiensis (Bt) crystal toxins genes, yielding 3-5% of the total leaf protein. Importantly, the insecticidal protein could be translated from the bacterial AU-rich mRNA, while for nuclear expression only synthetic GC-rich mRNAs could be used. [21] Since then, the Maliga laboratory developed chloroplast expression tools that yield 25% tetanus subunit vaccine [22] and >45% GFP in tobacco leaves. [23] Their current goal is expression of orally bioavailable recombinant proteins in tobacco and lettuce chloroplasts.

Awards and honors

Related Research Articles

<i>Arabidopsis thaliana</i> Model plant species in the family Brassicaceae

Arabidopsis thaliana, the thale cress, mouse-ear cress or arabidopsis, is a small plant from the mustard family (Brassicaceae), native to Eurasia and Africa. Commonly found along the shoulders of roads and in disturbed land, it is generally considered a weed.

<span class="mw-page-title-main">Symbiogenesis</span> Evolutionary theory holding that eukaryotic organelles evolved through symbiosis with prokaryotes

Symbiogenesis is the leading evolutionary theory of the origin of eukaryotic cells from prokaryotic organisms. The theory holds that mitochondria, plastids such as chloroplasts, and possibly other organelles of eukaryotic cells are descended from formerly free-living prokaryotes taken one inside the other in endosymbiosis. Mitochondria appear to be phylogenetically related to Rickettsiales bacteria, while chloroplasts are thought to be related to cyanobacteria.

<span class="mw-page-title-main">Plastid</span> Plant cell organelles that perform photosynthesis and store starch

A plastid is a membrane-bound organelle found in the cells of plants, algae, and some other eukaryotic organisms. They are considered to be intracellular endosymbiotic cyanobacteria.

<span class="mw-page-title-main">Horizontal gene transfer</span> Transfer of genes from unrelated organisms

Horizontal gene transfer (HGT) or lateral gene transfer (LGT) is the movement of genetic material between organisms other than by the ("vertical") transmission of DNA from parent to offspring (reproduction). HGT is an important factor in the evolution of many organisms. HGT is influencing scientific understanding of higher-order evolution while more significantly shifting perspectives on bacterial evolution.

<span class="mw-page-title-main">Genetic transformation</span> Genetic alteration of a cell by uptake of genetic material from the environment

In molecular biology and genetics, transformation is the genetic alteration of a cell resulting from the direct uptake and incorporation of exogenous genetic material from its surroundings through the cell membrane(s). For transformation to take place, the recipient bacterium must be in a state of competence, which might occur in nature as a time-limited response to environmental conditions such as starvation and cell density, and may also be induced in a laboratory.

<i>Agrobacterium</i> Genus of bacteria

Agrobacterium is a genus of Gram-negative bacteria established by H. J. Conn that uses horizontal gene transfer to cause tumors in plants. Agrobacterium tumefaciens is the most commonly studied species in this genus. Agrobacterium is well known for its ability to transfer DNA between itself and plants, and for this reason it has become an important tool for genetic engineering.

<span class="mw-page-title-main">Gene gun</span> Device used in genetic engineering

In genetic engineering, a gene gun or biolistic particle delivery system is a device used to deliver exogenous DNA (transgenes), RNA, or protein to cells. By coating particles of a heavy metal with a gene of interest and firing these micro-projectiles into cells using mechanical force, an integration of desired genetic information can be introduced into desired cells. The technique involved with such micro-projectile delivery of DNA is often referred to as biolistics, short for "biological ballistics".

<span class="mw-page-title-main">Nuclear gene</span> Gene located in the cell nucleus of a eukaryote

A nuclear gene is a gene that has its DNA nucleotide sequence physically situated within the cell nucleus of a eukaryotic organism. This term is employed to differentiate nuclear genes, which are located in the cell nucleus, from genes that are found in mitochondria or chloroplasts. The vast majority of genes in eukaryotes are nuclear.

Extranuclear inheritance or cytoplasmic inheritance is the transmission of genes that occur outside the nucleus. It is found in most eukaryotes and is commonly known to occur in cytoplasmic organelles such as mitochondria and chloroplasts or from cellular parasites like viruses or bacteria.

<span class="mw-page-title-main">Gene delivery</span> Introduction of foreign genetic material into host cells

Gene delivery is the process of introducing foreign genetic material, such as DNA or RNA, into host cells. Gene delivery must reach the genome of the host cell to induce gene expression. Successful gene delivery requires the foreign gene delivery to remain stable within the host cell and can either integrate into the genome or replicate independently of it. This requires foreign DNA to be synthesized as part of a vector, which is designed to enter the desired host cell and deliver the transgene to that cell's genome. Vectors utilized as the method for gene delivery can be divided into two categories, recombinant viruses and synthetic vectors.

<span class="mw-page-title-main">Agroinfiltration</span> Method in plant biotechnology

Agroinfiltration is a method used in plant biology and especially lately in plant biotechnology to induce transient expression of genes in a plant, or isolated leaves from a plant, or even in cultures of plant cells, in order to produce a desired protein. In the method, a suspension of Agrobacterium tumefaciens is introduced into a plant leaf by direct injection or by vacuum infiltration, or brought into association with plant cells immobilised on a porous support, whereafter the bacteria transfer the desired gene into the plant cells via transfer of T-DNA. The main benefit of agroinfiltration when compared to the more traditional plant transformation is speed and convenience, although yields of the recombinant protein are generally also higher and more consistent.

<span class="mw-page-title-main">Transplastomic plant</span>

A transplastomic plant is a genetically modified plant in which genes are inactivated, modified or new foreign genes are inserted into the DNA of plastids like the chloroplast instead of nuclear DNA.

A transfer DNA (T-DNA) binary system is a pair of plasmids consisting of a T-DNA binary vector and a virhelper plasmid. The two plasmids are used together to produce genetically modified plants. They are artificial vectors that have been derived from the naturally occurring Ti plasmid found in bacterial species of the genus Agrobacterium, such as A. tumefaciens. The binary vector is a shuttle vector, so-called because it is able to replicate in multiple hosts.

<span class="mw-page-title-main">Plant genetics</span> Study of genes and heredity in plants

Plant genetics is the study of genes, genetic variation, and heredity specifically in plants. It is generally considered a field of biology and botany, but intersects frequently with many other life sciences and is strongly linked with the study of information systems. Plant genetics is similar in many ways to animal genetics but differs in a few key areas.

<span class="mw-page-title-main">Chloroplast DNA</span> DNA located in cellular organelles called chloroplasts

Chloroplast DNA (cpDNA) is the DNA located in chloroplasts, which are photosynthetic organelles located within the cells of some eukaryotic organisms. Chloroplasts, like other types of plastid, contain a genome separate from that in the cell nucleus. The existence of chloroplast DNA was identified biochemically in 1959, and confirmed by electron microscopy in 1962. The discoveries that the chloroplast contains ribosomes and performs protein synthesis revealed that the chloroplast is genetically semi-autonomous. The first complete chloroplast genome sequences were published in 1986, Nicotiana tabacum (tobacco) by Sugiura and colleagues and Marchantia polymorpha (liverwort) by Ozeki et al. Since then, a great number of chloroplast DNAs from various species have been sequenced.

<span class="mw-page-title-main">History of genetic engineering</span>

Genetic engineering is the science of manipulating genetic material of an organism. The first artificial genetic modification accomplished using biotechnology was transgenesis, the process of transferring genes from one organism to another, first accomplished by Herbert Boyer and Stanley Cohen in 1973. It was the result of a series of advancements in techniques that allowed the direct modification of the genome. Important advances included the discovery of restriction enzymes and DNA ligases, the ability to design plasmids and technologies like polymerase chain reaction and sequencing. Transformation of the DNA into a host organism was accomplished with the invention of biolistics, Agrobacterium-mediated recombination and microinjection. The first genetically modified animal was a mouse created in 1974 by Rudolf Jaenisch. In 1976 the technology was commercialised, with the advent of genetically modified bacteria that produced somatostatin, followed by insulin in 1978. In 1983 an antibiotic resistant gene was inserted into tobacco, leading to the first genetically engineered plant. Advances followed that allowed scientists to manipulate and add genes to a variety of different organisms and induce a range of different effects. Plants were first commercialized with virus resistant tobacco released in China in 1992. The first genetically modified food was the Flavr Savr tomato marketed in 1994. By 2010, 29 countries had planted commercialized biotech crops. In 2000 a paper published in Science introduced golden rice, the first food developed with increased nutrient value.

<span class="mw-page-title-main">Genetic engineering techniques</span> Methods used to change the DNA of organisms

Genetic engineering techniques allow the modification of animal and plant genomes. Techniques have been devised to insert, delete, and modify DNA at multiple levels, ranging from a specific base pair in a specific gene to entire genes. There are a number of steps that are followed before a genetically modified organism (GMO) is created. Genetic engineers must first choose what gene they wish to insert, modify, or delete. The gene must then be isolated and incorporated, along with other genetic elements, into a suitable vector. This vector is then used to insert the gene into the host genome, creating a transgenic or edited organism.

A plastid is a membrane-bound organelle found in plants, algae and other eukaryotic organisms that contribute to the production of pigment molecules. Most plastids are photosynthetic, thus leading to color production and energy storage or production. There are many types of plastids in plants alone, but all plastids can be separated based on the number of times they have undergone endosymbiotic events. Currently there are three types of plastids; primary, secondary and tertiary. Endosymbiosis is reputed to have led to the evolution of eukaryotic organisms today, although the timeline is highly debated.

<span class="mw-page-title-main">Maureen Hanson</span> American molecular biologist

Maureen Hanson is an American molecular biologist and Liberty Hyde Bailey Professor in the Department of Molecular Biology and Genetics at Cornell University in Ithaca, New York. She is a joint member of the Section of Plant Biology and Director of the Center for Enervating Neuroimmune Disease. Her research concerns gene expression in chloroplasts and mitochondria, photosynthesis, and the molecular basis of the disease Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS).

References

  1. Maliga, P, Sz-Breznovits, A, Marton, L (July 1973). "Streptomycin resistant plants from callus culture of haploid tobacco". Nature New Biology. 244 (131): 29–30. doi:10.1038/NEWBIO244029A0. PMID   4515911. S2CID   26838425.
  2. Cseplo, A, Maliga, P (November 1982). "Lincomycin resistance, a new type of maternally inherited mutation in Nicotiana plumbaginifolia". Current Genetics. 6 (2): 105–109. doi:10.1007/BF00435208. PMID   24186475. S2CID   7925902.
  3. Cseplo, A, Maliga, P (1984). "Large scale isolation of maternally inherited lincomycin resistance mutations in diploid Nicotiana plumbaginifolia protoplast cultures". Molecular and General Genetics. 196 (3): 407–412. doi:10.1007/BF00436187. S2CID   24421287.
  4. Svab Z, Maliga P (August 1991). "Mutation proximal to the tRNA binding region of the Nicotiana plastid 16S rRNA confers resistance to spectinomycin". Molecular and General Genetics. 228 (1–2): 316–319. doi:10.1007/BF00282483. PMID   1832206. S2CID   34949950.
  5. Cseplo, A, Medgyesy, P, Hideg, E, Demeter, S, Marton, L, Maliga, P (August 1985). "Triazine-resistant Nicotiana mutants from photomixotrophic cell cultures". Molecular and General Genetics. 200 (3): 508–510. doi:10.1007/BF00425742. S2CID   44212543.
  6. Moll, B, Polsby, L, Maliga, P (December 1989). "Streptomycin and lincomycin resistances are selective plastid markers in cultured Nicotiana cells". Molecular and General Genetics. 221 (2): 245–250. doi:10.1007/BF00261727. S2CID   19879921.
  7. Maliga, P (June 2004). "Plastid transformation in higher plants". Annual Review of Plant Biology. 55: 289–313. doi:10.1146/ANNUREV.ARPLANT.55.031903.141633. PMID   15377222. S2CID   36725756.
  8. Medgyesy, P, Fejes, E, Maliga, P (October 1985). "Interspecific chloroplast recombination in a Nicotiana somatic hybrid". Proceedings of the National Academy of Sciences of the United States of America. 82 (20): 6960–6964. Bibcode:1985PNAS...82.6960M. doi:10.1073/PNAS.82.20.6960. PMC   391289 . PMID   16593619. S2CID   20080861.
  9. Fejes, E, Maliga, P (1990). "Extensive homologous chloroplast DNA recombination in the pt14 Nicotiana somatic hybrid". Theoretical and Applied Genetics. 79 (1): 28–32. doi:10.1007/BF00223782. PMID   24226115. S2CID   1575818.
  10. Svab, Z, Hajdukiewicz, P, Maliga, P (1990). "Stable transformation of plastids in higher plants". Proceedings of the National Academy of Sciences of the United States of America. 87 (21): 8526–8530. Bibcode:1990PNAS...87.8526S. doi:10.1073/PNAS.87.21.8526. PMC   54989 . PMID   11607112. S2CID   8024062.
  11. Svab, Z, Maliga, P (February 1993). "High-frequency plastid transformation in tobacco by selection for a chimeric aadA gene". Proceedings of the National Academy of Sciences of the United States of America. 90 (3): 913–917. Bibcode:1993PNAS...90..913S. doi: 10.1073/pnas.90.3.913 . PMC   45780 . PMID   8381537. S2CID   7281078.
  12. Carrer, H, Hockenberry, TN, Svab, Z, Maliga, P (October 2004). "Kanamycin resistance as a selectable marker for plastid transformation in tobacco". Molecular and General Genetics. 241 (1–2): 49–56. doi:10.1007/BF00280200. PMID   8232211. S2CID   2291268.
  13. "Step Seen Toward Altering Photosynthesis". New York Times. November 2, 1990.
  14. Yu, Q, Lutz, K, Maliga, P (2017). "Efficient plastid transformation in Arabidopsis". Plant Physiology. 175 (1): 186–193. doi:10.1104/pp.17.00857. PMC   5580780 . PMID   28739820. S2CID   206339557.
  15. Lutz, K, Maliga, P (2007). "Construction of marker-free transplastomic plants". Current Opinion in Biotechnology. 18 (2): 107–114. doi:10.1016/J.COPBIO.2007.02.003. PMID   17339108. S2CID   40899963.
  16. Hajdukiewicz P, Svab Z, Maliga P (September 1994). "The small, versatile pPZP family of Agrobacterium binary vectors for plant transformation". Plant Molecular Biology. 25 (6): 989–994. doi:10.1007/BF00014672. PMID   7919218. S2CID   9877624.
  17. Matsuoka, A, Maliga, P (2021). "Prospects for Reengineering Agrobacterium tumefaciens for T-DNA delivery to Chloroplasts". Plant Physiology. 186: 215–220. doi:10.1093/plphys/kiab081. PMC   8154051 . PMID   33620481. S2CID   232017220.
  18. Allison, LA, Simon, LD, Maliga, P (1996). "Deletion of rpoB reveals a second distinct transcription system in plastids of higher plants". The EMBO Journal. 15 (11): 2802–2809. doi:10.1002/j.1460-2075.1996.tb00640.x. PMC   450217 . PMID   8654377. S2CID   39505448.
  19. Hajdukiewicz, P, Allison, LA, Maliga, P (1997). "The two RNA polymerases encoded by the nuclear and the plastid compartments transcribe distinct groups of genes in tobacco plastids". The EMBO Journal. 16 (13): 4041–4048. doi:10.1093/emboj/16.13.4041. PMC   1170027 . PMID   9233813. S2CID   10769603.
  20. Suzuki, J, Ytterberg, AJ, Beardslee, TA, Allison, LA, vanWijk, KJ, Maliga, P (2004). "Affinity purification of the tobacco plastid RNA polymerase and in vitro reconstitution of the holoenzyme". Plant J. 40 (1): 164–172. doi: 10.1111/J.1365-313X.2004.02195.X . PMID   15361150. S2CID   24662704.
  21. McBride, KE, Svab, Z, Schaaf, DJ, Hogan, PS, Stalker, DM, Maliga, P (1995). "Amplification of a Chimeric Bacillus Gene in Chloroplasts Leads to an Extraordinary Level of an Insecticidal Protein in Tobacco". Bio/Technology. 13 (4): 362–365. doi:10.1038/NBT0495-362. PMID   9634777. S2CID   2154428.
  22. Tregoning, J, Nixon, P, Kuroda, H, Svab, Z, Clare, S, Bowe, F, Fairweather, N, Ytterberg, J, vanWijk, KJ, Dougan, G, Maliga, P (August 1985). "Expression of tetanus toxin fragment C in tobacco chloroplasts". Nucleic Acids Res. 31 (4): 1174–1179. doi:10.1093/NAR/GKG221. PMC   150239 . PMID   12582236. S2CID   15125002.
  23. Yu Q, Tungsuchat-Huang T, Verma K, Radler MR, Maliga P (August 1985). "Independent translation of ORFs in dicistronic operons, synthetic building blocks for polycistronic chloroplast gene expression". Plant J. 103 (6): 2318–2329. doi: 10.1111/tpj.14864 . PMID   32497322. S2CID   219328497.