Archaea Groups: Phylogeny and Function

• The text describes the phylogenetic and functional diversity of primary groups of Archaea and evidence that has contributed to our understanding of archaeal diversity.

Korarchaeota

• Uncultivated
• Hyperthermophile
• Originally defined phylogenetically

Nanoarchaeota

• Obligate symbionts
• Small genomes

Euryarchaeota

• Thermophiles/hyperthermophiles
• Extreme halophiles
• Methanogens
• Non methanogens: Thermoplasma/Ferroplasma
• Acidophiles: obtained from coal refuse piles
• Marine Group II Euryarchaeota
• Relatives of Thermoplasma
• Live in the open ocean; quantified planktonic archaea in the ocean by rDNA hybridization

Crenarchaeota and Thaumarchaeota

• First discovered isolates were extremophiles (thermophiles/hyperthermophiles)
• Mesophiles also
• Much Crenarchaeota diversity is uncultivated
• Sulfolobus solfataricus lives in hot, sulfur-rich environments.
• Pyrolobus fumarii lives in walls of black smokers (hydrothermal vents) with the highest temperature for life (113°C)
• Marine Group I Crenarchaeota
• Thermophile relatives
• Live in open ocean
• Thaumarchaeota are part of Crenarchaeota
• Ammonia-oxidizing archaea may be present in Thaumarchaeota

Discovery of Ammonia-Oxidizing Archaea (AOA)

• 16S rRNA surveys revealed abundant archaeal sequences.
• Gene homologs of bacterial amoA (ammonia monooxygenase) were identified in archaea genomes.
• Nitrosopumilis maritimus was the first cultivated AOA, performing autotrophic ammonia oxidation (nitrification).
• Cenarchaeum symbiosum is an extracellular ammonia-oxidizing symbiont of sponge (Axinella mexicana).
• AOA are more abundant than AOB and are more efficient at low ammonia concentrations.

Darwinian Threshold

• Described in the context of cellular and microbial evolution
• Represents the transition from precellular life to cellular life in LUCA.
• DNA replication, transcription, and translation cross the Darwinian threshold first
• Transition from mostly horizontal gene transfer (HGT) to vertical gene transfer.

Molecular/Cellular Similarities Between Archaea and Eukaryotes

• Ribosome for translation is shared between all domains of life
• Homologs exist between archaea and eukaryotes including:
• Transcription machinery
• ATP synthase
• DNA polymerase

Lipid Divide

• The uncertainty around what the common ancestor of bacteria, eukaryotes, and archaea was like
• Bacteria and eukaryotes: ester linkages
• Archaea: ether linkages
• Evolution of membranes leads to compartmentalization of cellular processes and regulation of what goes in and out of the cell.

Modern Challenges of Molecular Systematics and Classification of Microbial Eukaryotes

• Issues with rRNA phylogenies.
• Morphology may not always align with molecular systematics.
• Discrepancies exist between phylogenetic markers, like rRNA vs. other genes.
• The chimeric nature of eukaryotic genomes, a mix of organisms (especially for symbionts)
• Lack of bootstrap support for supergroups.

Asgard Archaea Identification

• Discovered in deep-sea sediments with MAGs
• Eukaryotic signature proteins (ESPs) found only in eukaryotes were discovered in the Asgards.
• Closest known relatives of eukaryotes.
• Exhibit more metabolic diversity than eukaryotes.

Roles of Microbial Eukaryotes in Carbon Cycle

• Encompasses both aerobic and anaerobic roles.
• Photosynthesizers fix carbon and turn it into rock; protists form shells, sink into the sediment, and become fossil carbon.
• Predators
• Heterotrophs

"SAR" Group

• Refers to Stramenopiles, Alveolates, and Rhizaria and is the most abundant group
• Stramenopiles
• Characterized by hairy flagella
• Composed of mix of phototrophs and heterotrophs
• Examples: diatoms (phototrophs with shells), kelp (brown algae), and oomycetes
• Alveolates
• Primarily parasites of humans and other organisms
• Feature sacs (alveoli) underneath the outer cell membrane
• Examples: dinoflagellates, apicomplexa, and ciliates
• Rhizaria have some version of a shell
• Example: amoeba

Anaerobic Eukaryotes vs. Aerobic Eukaryotes

• Anaerobic eukaryotes
• Cannot do aerobic respiration, relying only on fermentation/SLP
• Have atypical or no mitochondria
• Often have hydrogenosomes

Primary Supergroups of Eukaryotes

• Opisthokonts: Animals + fungi + choanoflagellates
• Amoebozoa
• Rely on amoeboid forms
• Use actin-based motility
• Types of multicellularity
• Cohesive (occurs in humans)
• Develops from a single progenitor cell, clonal division, physically attached organisms
• Long-term multicellular body w/ specialized cells
• Aggregative
• Cells come together and temporarily aggregate in response to environmental cues/starvation and may separate later
• Individual to multicellular/syncytial forms differentiate into fruiting bodies

Mechanisms of Bacterial-Eukaryote Symbiosis

• Includes primary, secondary, and tertiary symbioses; important in organelle evolution and the creation of diversity of eukaryote groups.
• Endosymbioses are common, but genetic fixation of them into organelles is rare
• Example: nitroplast, a nitrogen-fixing organelle originally a co-evolved endosymbiont in marine alga
• Shapes the diversity of eukaryotes and provides novel characteristics
• Symbionts have reduced genomes but control their replication, division, and expression
• Organelles have only essential genes
• Division is controlled by the cell
• The number of membranes indicates the number of endosymbiosis events
• Primary endosymbiosis: two membranes; host cell engulfs bacterium, bacterial membrane is retained
• Secondary endosymbiosis: three membranes; host engulfs cell with primary endosymbiont
• Tertiary endosymbiosis: four membranes; host engulfs cell with secondary endosymbiont

Hydrogenosomes

• Eukaryotes do not necessarily all have mitochondria so don't always need to use O₂
• Used for fermentation/SLP
• Often have hydrogenosomes, double membrane-bound organelles, that generate H2/CO2 and acetate (1 ATP)
• Production of H₂ and CO2 is not energetically favorable
• Interspecies hydrogen transfer (syntrophy with methanogens) pulls the hydrogenosome reaction forward
• Hydrogenosomes may have originated from mitochondrial reduction
• Mitochondria → anaerobic mitochondria (use alternative electron acceptor)
• Anaerobic mitochondria → hydrogenosome

Mobile Genetic Elements (MGEs)

• MGEs transfer genetic material from one host to another
• Genetic material also persists within the host
• Plasmids (transformation/conjugation)
• Can be chimeric (mixtures of genomes from different hosts
• Encode genes for their own transfer and antibiotic resistance
• Conjugative plasmids: self-transfer due to conjugation (tra) genes, have oriT sequences, and often encode antibiotic resistance genes (ARGs)
• Mobilizable plasmids: do not encode genes for self-transmission but can have ARGs, can be transferred if another plasmid in the same cell has conjugation genes
• "Piggybacking” on another plasmid
• Viruses: transduction
• Transposons: transposition
• Integrative and conjugative elements: conjugation
• Can cross with transposons

Conjugation, Transduction, and Transformation

• Conjugation involves cell-to-cell transfer of bacterial genes via physical interaction between cells.
• Transduction involves phage-mediated transfer of bacterial genes.
• Transformation involves the bacterial cell taking up genetic material from the environment (e.g., DNA released when another cell lyses).

Plasmid Replication Mechanism

• Relies on “rolling circle replication”
• Ori is the origin of replication in the plasmid
• Generally high in As and Ts
• Uses both host machinery and plasmid rep proteins
• RepA is an initiator protein that nicks one strand
• Host proteins used: ligase, single-stranded binding proteins
• Steps of rolling circle replication
• RepA nicks DNA at the origin cut site
• A new strand is synthesized, continues in a circle
• The new strand is cleaved and ligated

"Accessory" Genes on Plasmids

• Toxins/colicins, antibiotics, and cognate resistance genes; the producer is immune
• Virulence genes
• Degradative enzymes

Structural and Genetic Components of Phages

• Structural
• Capsid
• Tail
• Proteases
• Genetic components
• Lots of accessory genes

Bacteriophage Life Cycles

• Lytic: new viral particles are made, the host cell lyses and releases progeny viruses, horizontal transmission of the phage genome occurs
• Lysogenic: virus integrates into the host genome, no production of new viral particles
• a cell becomes a lysogen
• integrated phage = prophage
• replicated along with the host genome, vertical transmission of phage genome
• induction: activation of prophage to the lytic cycle

Host Cell "Barriers" to Horizontal Gene Transfer

• Successful infection
• Modification of virus receptor
• Degradation by restriction enzymes
• Modification of viral DNA preventing replication
• Abortive infection: host cell commits suicide
• CRISPR Cas system recognizes foreign DNA and degrades it

Host Cell Defenses

• Host-encoded restriction enzymes
• RE cuts viral DNA
• Host also encodes a methylase that protects its own DNA
• CRISPR-Cas9
• CRISPR spacers store records of prior infections by phage
• Adaptive immunity in bacteria
• Heritable; a cell does not have to be infected to be immune to that phage
• Matches a phage DNA sequence, targeting a specific part of the phage genome

Giant Viruses and Virophages

• Giant viruses
• Infect eukaryotes
• Get engulfed/eaten (phagocytosis)
• Can reprogram eukaryotic hosts, which might give some competitive advantage
• Giant size, large DNA genomes, have translation components, assemble in viral factories.
• Virophage
• A virus that can infect other viruses, usually giant viruses
• Can be integrated into the host genome and defend against other viruses which increases host cell survival

Transposons

• Transposons are used for bacterial mutagenesis
• These contain terminal inverted repeats, when inserted, flanking direct repeats occur
• Transposase is the enzyme that mobilizes transposons (often encoded by transposon)
• Conservative transposition (cut and paste)
• A transposon is excised from donor DNA then inserted in different genome
• Replicative transposition (copy and paste)
• The transposon is replicated keeping the donor strand unharmed
• Transposon-based mutagenesis (random)
• Transposons are added to create mutants
• Can select for different mutant phenotypes
• Conjugative transposons
• carry genes that enable conjugation
• excision from donor DNA → conjugative transfer to recipient cell; integrates into the recipient cell DNA
• enable both mobilizable plasmids and non-conjugative transposons to be transferred via conjugation

Spore Formation

• Endospores
• Endospore formation only in Firmicutes (Bacillus and Clostridium)
• Bacillus thuringiensis: endospore coat contains insecticidal proteins, used for genetic engineering
• Sporulation: sensing of conditions, nutrient deprivation, and population density.
• Cells are committed once sporulation starts, as signals accumulate
• Chromosome stretches out → asymmetric cell division begins and generate mother cell + forespore cells that eventually die and release more spores
• Streptomyces Spores
• Streptomyces grow in filaments
• Older filaments lyse as nutrients run out, secreting antibiotics
• Aerial mycelia grow and produce spores at their tips
• --Metabolically active, not as resistant as endospores
• Myxospores -- Formed by myxobacteria upon starvation, aggregate and form fruiting bodies that contain myxospores (not as tough as endospores)

Metabacterium Polyspora and Epulopiscium

• Metabacterium polyspora
• Reproduces via formation of multiple endospores
• Endospore former in Firmicutes, inhabits guinea pig GI tract
• Sporulation is tied to the timing of passage through the GI tract
• Guinea pig ingests feces w/ spores, which germinate in the upper intestine
• Spore formation = primary reproductive mechanism, part of normal life cycle
• Forespores are formed at poles and can do binary fission
• Epulopiscium
• Reproduces via production of multiple live offspring
• Mother cell lyses

Caulobacter

• Cell division via unequal binary fission and dimorphic life cycle
• Motile swarmer cell is chemotactic, used for dispersal, does not replicate and can't go back to stalked cell
• Stalked cell can divide and make new swarmer cells
• length is inversely correlated with nutrient concentration, stalk has more surface area for nutrient uptake

Bacterial Motility

• Requires motility apparatus and sensory system that relies on chemical gradients
• Swimming motility
• Flagellar or non-flagellar
• Bacterial flagellum: helical rotary structure, external, powered by PMF; flagellum proteins self-assemble, do not want to make until outside the cell.
• Start w/ basal body and rod, then hook assembly, then create filament
• Flagella often upregulated in nutrient-poor conditions
• Surface Motility
• Twitching motility with type IV pili (Pseudomonas aeruginosa)
• powered by ATP
• Myxococcus Motility
• Adventurous (A-motility): form of gliding motility, move independently by anchoring and retraction
• Social (swarms)
• Relies on type IV pili

Similarities and Differences Between Bacterial and Archaeal Mechanisms of Swimming Motility

• Bacterial flagellar motility acts as a corkscrew propeller
• CCW rotation → movement
• CW rotation → tumbling (random)
• Archaeal swimming motility
• Archaeal flagella (archaellum): components are homologous to type IV pili
• Rotate as bundles ("tufts" of flagella)
• Some do not extend and retract
• Chemotaxis system proteins (e.g. MCPs) are conserved in bacteria and archaea
• Haloarchaea are some motile and phototactic using sensory rhodopsins to sense different wavelengths of light

Spirochete Flagellar Motility

• Flagella located in periplasm: "endoflagella“ wrapped around the cell body
• Flagella at each end must be rotating in opposite directions
• Same direction = flexing (similar to tumbling)
• Advantages: allows for movement in viscous liquids, flagella are protected, internal flagella do not elicit an antigenic response

Bacterial Chemotaxis

• Switches between run and tumble, the frequency allows for chemotaxis
• If moving toward attractant: extend runs
• Machinery
• Attractants = nutrients
• Repellants = toxic compounds
• Chemosensory array at one end of the cell
• Methyl-accepting chemotaxis proteins (MCPs) = sensor (sensory domains, output domains) that monitors sensitivity based on methylation
• CheA = histidine kinase; integrates signal
• CheY = response regulator (flagellar motility); takes the input and directs it to the response
• With the use of bacterial methyl-accepting chemotaxis proteins in the process, magnetotaxis can increase the efficiency of aerotaxis as well
• Cells are pulled into alignment by a magnetic field (passive, does not expend energy)
• Magnetosomes are membrane-enclosed structures containing magnetic mineral (compass)

Quorum Sensing

• Cell density-dependent regulation that allows bacteria to monitor their population and respond accordingly
• Used for interspecies communication; autoinducers are shared across species
• Bacterial luminescence
• Once population threshold is reached, the switch for luminescence is flipped and chemical energy is changed into light energy
• Aliivibri fischeri in symbiosis with Hawaiian bobtail squid -Aliivibri gets protection and nutrient supply
• • Squid development depends on* colonization (Vibrio alters host gene expression) - host receptors detect specific symbiont chemicals and make specific antimicrobials to get rid of other potential colonizers
  • Luminescence genes are transferable to other species
• Pathogenesis (Pseudomonas aeruginosa)
• Forms biofilms on surfaces protected by exopolysaccharide matrix
• Resistant to antibiotic treatment- -uses quorum sensing or biofilm production, and then virulence factor production with the Las system and Rhl system
• Las system controls Rhl system -- Las system produces second autoinducer for Rhl system
• Can target quorum sensing signals for treatment therapies

Bacterial Luminescence and Quorum Sensing

• In bacterial luminescence, quorum sensing plays a role, and LuxR and LuxI proteins have various functions
• Genes for luminescence are carefully controlled given their huge energy demand of 50 ATP for one photo of light
• Quorum sensing involves detecting a diffusible small molecule (autoinducer or pheromone) as it increases in concentration
• Enough cells= enough signal to produce active transcription
• LuxI protein synthesizes autoinducer and the autoinducer diffuses into the medium and accumulates
• At threshold concentration, autoinducer diffuses back into the cell and binds LuxR to promote lux transcription (light production)