Week 4 => Organelles and their genomes Flashcards
What are organelles?
While most cell biologists consider the term organelle to be synonymous with cell compartment, a space often bound by one or two lipid bilayers, some cell biologists choose to limit the term to include ONLY THEOSE CELL COMPRTMANETS THAT CONTAING DNA, having originated from formerly autonomous microscopic organisms acquired via ENDOSYMBIOSIS
Where are organelles found?
Cytosol (fluid that contains organelles; with which, comprises cytoplasm)
Why comparing what sequence in Mt DNA of various eukaryotes, various bacteria, and archaea where the origins of mitochondria narrowed down to?
Ribosomal RNA gene
What two bacterial endosymbionts are hypothesized to the source of mitochondria based on their 16S ribosomal RNA sequence?
Agrobacterium tumefaciens and Pseudomonas testosteroni
What do Agrobacterium tumefaciens and Pseudomonas testosteroni represent of the so-called purple bacteria?
the alpha and beta subdivision respectively
What is alphaproteobacteria?
Agrobacterium tumefaciens
Which endosymbiont subdivision have rise to mitochondria?
The alpha subdivision, a group that also contains the rhizobacteria, the agrobacteria, and the rickettsia’s - all the prokaryotes that have developed intracellular or other close relationships with eukaryotic cells
Rickettsia Prowazekii (R. prowazekki)
- an aerobic intracellular pathogen
- Causative agent of epidemic typhus
- Genome is approximately 1.1 MBp and has 834 protein genes
- Intriguing similarities with mitochondria (e.g., AT-biased, reduced genome), but nevertheless clearly the product of independent reduction evolution
- Decedent of alphaproteobacterial lineage
Mitochondrial genomes can be:
- circular
- circular mapping
- linear with short (3-100 bp) terminal repeats (triangles and diamonds)
The conventional multipart model of mitochondria genome
Where recombination between large repeats interconverts between a master circle conformation and a set of sub genomic circles
Alternative physical structure of mitochondrial genome that can all produce circular genomes?
- Circular chromosome
- Head-to-tail concatemers
- Circularly permutated linear molecules
What have gene-poor, compact mitochondria genomes?
Animals, fungi, and many single-celled eukaryotes
Homo sapiens mtDNA
- 16569 bp
- non-stranded genetic code (UGA as tryptophan, AUA as methionine, not isoleucine)
- 13 protein genes (All part of electron transport chain)
- 22 tRNA
Plasmodium falciparum (malaria parasite) mitochondrial genome
- 5967 bps
- stranded genetic code
- 3 protein genes
- fragmented rRNA genes
- no tRNA genes (tRNA must be imported from host)
What has gene-rich mitochondrial genomes?
Jakobid flagellates
Example of Jakobid flagellates bacteria
Reclinomonas americana, a freshwater protist
Reclinomonas americana mtDNA key features
- Reclinomas and its relatives have the most gene-rich mtDNAs known
- 100 genes (approximetly 65 protein genes)
- Shine-Dalgarno (ribosome-binding) sequences
- genome encodes RNA component of RNAse P (a tRNA processing complex)
- strikingly bacterial four-subunit RNA polymerase (all other eukaryotes have a single-subunit phage-type RNA polymerase)
What have bloated mitochondrial gneomes?
Plants
Bloated mitochondrial genomes
- 1685 kb mitochondrial genomes of cucumber consisting of THREE circular-mapping chromosomes
- large size result of proliferation of repeats, introns and acquisition of genes from nuclear and chloroplast genomes, viruses, and bacteria
- Chloroplast sequences include functional tRNA genes
What may explain apparent unidirectional plasmid-to-mitochondrion gene transfer?
Mitochondria are known to have a \DNA uptake system, while chloroplasts (plasmids) do not
Mitochondrial genome size vs coding capacity
Non-coding DNA accounts for most of the genome size variation seen in mitochondria (thus larger size more non-coding DNA)
Mitochondrion-related organelles (MROs)
Reduced, simplified “remnants” of mitochondria
Hydrogenosomes
- an MRO
- Peculiar eukaryotic organelles that generate ATP and evolve H2 gas
- Have evolved (from mitochondria) multiple times independently
- Some have a small genome, other have no genome at all
- some enzymes involved in anaerobic hydrogenesomal biochemistry appear to have been acquired from anaerobic bacteria by LGT
- Site of iron-sulfate cluster biogenesis (reason they persist)
Mitosome
- Mitosomes are mitochondrion-related organelles found in a range of unicellular eukaryotic organisms that inhabit oxygen-poor environments.
- MROs
Example of diplomonad
Giardia lamblia
Giardia lamblia
- The only known function of mitosomes in Giardia is iron-sulfate cluster biosynthesis
- Mitosomes no longer make ATP; synthesis of ATP takes place in cytosol
- like hydrogenosome-containig organisms, LGT from prokaryotes appears to have played a role in how mitosome-bearing eukaryotes have adapted to anaerobic environments
Monocernomonoides sp.
An anaerobic protist living in the guts of chinchillas
Complete mitochondrial loss
Mitochondrial loss enable by LGT of four genes encoding enzymes for SUF-type iron-sulfur cluster biosynthesis pathway that functions in the cytosol, not in the mitochondria (like other eukaryotes)
IscU proteins
Iron-sulfur cluster assembly enzyme ISCU, mitochondrial is a protein that in humans is encoded by the ISCU gene.
IscS proteins
a cysteine desulfurase that is proposed to sequester inorganic sulfur for Fe–S cluster assembly
What do Hsp60 and Hsp70 protein transport bring into the mitochondria?
IscU and IscS for iron-sulfate cluster biosynthesis
How do we know that hydrogenosomes and mitosomes are mitochondrion-related organelles?
The nuclear genomes of these organisms encode obvious MRO-targeted proteins AND the protein import apparatus is demonstrably mitochondrial (i.e., the mechanisms of protein import is conserved amongst all mitochondria and mitochondrion-derived organelles)
Significance of cyanobacterium and eukaryotes endosymbiosis?
Group of prokaryotes that “invented” oxygenic photosynthesis
What are the three branches eukaryotes with cyanobacterium evolve to?
- Red algae
- Green algae
- Glaucophytes
Primary endosymbiosis
Primary endosymbiosis is the process by which a eukaryotic cell engulfs a prokaryotic cell, creating organelles like mitochondria and chloroplasts. It is considered the first endosymbiotic event.
Chloroplast primary endosymbiosis
Eukaryotes with prokaryote (cyanobacterium) leading to oxygen photosynthetic organissm
Plasmid evolution
Involved Endosymbiotic Gene Transfer (EGT) and the development of a protein import apparatus
What did the primary plasmid-containing cell originate from?
Primary endosymbiosis event of non-photosynthetic eukaryote and cyanobacteria
TOMs
Translocons of the outer mitochondrial membrane
TIMs
Translocons of the inner mitochondrial membrane
TOCs
Translocons of the outer chloroplast membrane
TICs
translocons of the inner chloroplast membrane
Translocons
The translocon is a complex of proteins associated with the translocation of polypeptides across membranes
What mediates the translocation of proteins?
Protein machinery (TOMs/TOCs and TIMs/TICs) on the outer and inner membrane
What type of ‘signal sequences’ do nucleus-encoded protein have?
N-terminal (‘transit’ or “target’ peptides (TPs))
Transit peptides
- 10-100 amino acids in length
- Rich in hydrophobic and positively charged amino acids
- recognized by receptors on the outer surface of the organelle
What is removed once proteins are inside the organelle?
The transit peptide
What was ‘inserted’ ii eukaryotes during the primary and second ensosymbiosis?
- primary => cyanobacterium
- secondary => a second mitochondria, nucleus, and a plastid (chloroplast) => the primary endosymbiont
Two types of plastids
- Primary plastids
- Secondary plastids (also known as ‘complex’ plastids)
Primary plastids
- evolved directly from cyanobacteria
- surrounded by two membranes
- the primary plastid of glaucophyte algae, red algae, and green algae are probably the result of a single primary endosymbiotic event
Secondary Plastids
- Acquired indirectly from primary plastids-bearing algae
- Surrounded by 3 or 4 membranes, ‘nucleomorph’ sometimes present
- Evolved at least 3 times (involving both red and green algal endosymbiosis)
- Secondary plastids are IMPORTANT: some of the most abundant photosynthetic microbes on the planet acquired their plastids in this manner
Nucleomorphs
Nucleomorphs are small, vestigial eukaryotic nuclei found between the inner and outer pairs of membranes in certain plastids.
What contains nucleomorphs?
Certain secondary plastids
Plasmid genomes predominantly resemble ___ genomes
Prokayrotic
Features of most plastid genomes:
- Circular
- Genes on both strands (bacterial type operons clearly present)
- Inverted repeats (IRA and IRB)
- Large (LSC) and small (SSC) regions have single-copy (SC) genes
What encodes the ribosomal RNA operon in plasmid genomes?
Inverted repeats
______ are greatly reduced in size, but tend to retain the overall structure of their photosynthetic counterparts
Non-photosynthetic plastids
what comprises the largest and most divergent plastid genomes known?
The new red algal subphylum proteorhodophytina
Parasite plants
- Lost the capacity to carry out photosynthesis
- genomes reduced in size and gene content
Balanophora laxiflora
- Holoparasitic plant
- 19 protein-coding genes (highly reduced)
- A/T rich protein-coding (extreme composition bias 12.2% G/C)
- Alternative genetic code (UAG = Trp)
- no tRNA genes (must be imported into plastid)
Plastome
Plastid genome
All known plastid genomes are significantly ___ compared to cyanobacterial genomes
Reduced
What have the largest genomes among plastids?
Red algal and red-algal derived secondary plastids
What re plastid genome mostly comprised of?
Proteins involved in photosynthesis and translation (note a reducing in the ‘other’ functional category, which includes transcription)
Component of the light harvesting apparatus, electron transport chain, and ATP synthase complex are a mixture of ___
Nucleus - and plastid-encoded proteins
Stroma
Stroma is a type of tissue that provides structure and support to organs, glands, and other tissues in the body. It can also refer to the colorless fluid in the chloroplast of a plant cell.
Thylakoid lumen
The thylakoid lumen represents a continuous exoplasmic cellular space that is poorly characterized compared with the other chloroplast compartments. Interest in the lumenal side of the thylakoid membrane has mainly focused on electron transport events associated with the inner membrane surface.
Different signals required on Primary plasmids vs secondary plasmids?
- Primary has only Transit peptides (TP)
- Secondary has TP and signal peptides (SP)
Nucleomorph (NM) where they are found in nature
- Maintained a nucleus in the plasmid
- Found in some algae with secondary plastids
- Located between the inner and outer pairs of plastid membranes
Where does the nucleomorph reside in organisms?
Within the ‘periplastidial compartment’ (PPC), which is the former cytosol of the algal endosymbiont
Nucleomorphs by definition
Are residual nuclei of endosymbiotic origin found in cryptomonads and cholarachinophytes
Cryptomonad
- Red algal origin
- Nucleomorphs embedded in the ER
Chlorarachniophyte
- Green algal origin
- Nucleomorphs separated from the nucleus, but still has membrane it needs to pass though
Nucleomorphs genomes are ____
- Tiny (3 chromosomes)
- Dense (1 gne/Kbp) (numerous overlapping genes)
- Low G/C content (25%)
- ultra-fast-evolving genes (>30% ORFans)
What happens when different genomes are stuck together for a long time?
Endosymbiotic gene transfer (EGT)
Endosymbiotic gene transfer (EGT)
- The process by which fragments of endosymbiont or organellar DNA end up in the nuclear genome
- LGT
Comparative genomic: compare organellar and nuclear gnomes to those of bacteria
Learn about where mitochondria and plastids came from and the impact of endosymbiosis on the eukaryotic nuclear genome
Comparative genomics: compare organellar genomes from species A, B, and C
Compare them to the nuclear genomes from species A, B, and C etc. etc. Such comparison can provide clues as to how organellar DNA evolves and migrates to the nucleus over ‘shorter’ timescales
Experiential systems to study EGT in the lab
- Involve transgenic organisms to observe EGT in ‘real time’
- Can provide insight into the mechanisms and frequency of EGT (has been done with yeast and tobacco)
EGT takes place over different timescales
1) Ancient transfer => during evolution of organelles from their bacterial progenitors
2) somewhat ‘recent’ transfers => Functional gens giving rise to different organellar genomes size and lineages
3) Very recent transfers => Giving rise to non-functional DNA fragments
NORGs
Nuclear integrants of organellar DNA
NUMTs
Nuclear mitochondrial DNA - mitochondrial DNA fragments integrated into the nuclear genome
NUPTs
Nuclear plastid DNA - plastid DNA fragments integrated into the nuclear genome
NUNMs
Nuclear nucleomorph DNA - fragments of DNA from nucleomorph genomes integrated into the nuclear genome (limited to single-celled algae with nucleomorphs)
NUMTs causing human genetic disease (patient of Chernobyl accident), result of 72 Bp insertion of mtDNA into the nuclear gene GLI3?
Created a premature stop codon, (presumably) resulting in a non-functional protein
How is DNA made available for integration?
Organelle lysis
How is DNA integrated?
- Non-homologous end joining at sites of double-strand break (DSB) repair
- Integration appears to eb non-random: EGT into A+T rich, open chromatin regions is more common
- NOTE: cDNA can also be integrated, i.e., organellar RNA can serve as substrate for EGT
What is the fate of transferred DNA (and the donor DNA in the organelle)?
- The majority of EGTs are non functional: they become pseudogenes and disappear or degenerate due to mutations
- The donor DNA still exist in the organelle and so ‘no harm done’
How do transferred gene acquire gene expression and targeting elements?
They acquire them de novo, but they can also acquire them from pre-existing genes
The limited transfer window hypothesis
- There is a correlation between the number of organelles per cell and the frequency of EGT from that organelle
- for example: polyplastidic species have 80 times more NUPTs than monoplastidic species
- explains the persistence of nucleomorphs
endosymbiotic gene transfer
- Organelle-to-nucleus transfer is/has been FRQUENT
- Nucleus-to-organelle transfer is/has been RARE
If EGT is so important why hasn’t it gone to completion?
In some cases, it has (e.g. in some secondary plastid-bearing algae, whose nucleomorphs and nucleomorph genomes have disappeared completely). For the most part, however, mitochondria and plastids retain a genome
What are the most common interorganelle DNA transfer types?
- Plastid-to-nucleus
- Mitochondria-to-nucleus
Hypothesis for “Why do organelles retain genomes?”
- Hydrophobicity hypothesis
- Redox regulation hypothesis
- Genetic code disparity hypothesis
Why do organelles retain genomes?
There is no single answer: the reasons why organelles retain genomes may differ from gene to gene, organelle to organelle, and lineage to lineage
Hydrophobicity hypothesis
Protein encoded in organellar genomes are extremely hydrophobic, and this would be difficult/impossible to import across the membranes of the organelle should their genes reside int he nuclear genome
Redox regulation hypothesis
The expression of key proteins encoded in organellar genomes, in particular those involved in electron transport and energy coupling, must be quickly and directly regulated by the redox state of the mitochondria. It is important for the cell to have the genes for such proteins located ‘on site’ - in the same compartment in which their proteins actually function
Genetic code disparity hypothesis
All animals and most fungi have at least one genetic code disparity between their nuclear and mitochondria genomes (this would render functional gene transfer impossible)
Organelle
- A specialized part of the cell having specific function; a cell organ
- A structure or part that is enclosed within its own membrane inside a cell and has a particualr function
- found only in eukaryotic cells and are absent from the cells of prokaryotes life bacteria
Paulinella chromatophora
- since-celled eukaryote, in the the Rhine river
- 1 or 2 green-pigmented ‘chromatophores’ per cell that divided synchronously with the host cell
- This symbiosis is 60 million years old according the ‘molecular clock’ estimates
Chromatophore genome: key features
- 1.02 megabase pairs in size, 867 poteen-coding genes
- High degree of gene order conservation between chromatophore and cyanobacterium WHY5701, but only 1/3 the size
- Many pseudogenes present (undergoing reductive evolution)
What is a recurring theme in organelle biology and evolution?
The use of N-terminal extensions as mediators of protein trafficking
