Practical Phylogeny

Practical Phylogeny

An increasingly serious bottleneck in molecular biology arises from the relative ease of determining a human gene sequence and the relative difficulty of determining its function. I propose here a systematic tool that addresses this problem in the situation where the new gene does not seem to have known homologues in better studied lower organisms such as nematode, fruit fly, yeast, and bacteria.

The idea is to assign the human gene to its homologous superfamily as quickly as possible, by bridging the gap to targeted organisms with the least amount of sequencing. This is done by identifying key intermediate species in the phylogenetic tree and sequencing the genes there, until a reliable homology probe is developed that can reliably pick out the homologues in organisms having more rapid and less expensive experimental milieux.

The reason for this is that open reading frames have been determined for all yeast genes and very rapic progress is being made in assigning them functions, structural domains, and substrate ligands. Once the unknown human gene is linked into yeast, it is immediately linked into every other organism for which something is known about the gene family. The nematode, C. elegans, forms a nearer intermediat to humans than yeast -- it will be completely sequenced by 1998. Tapping into a gene's superfamily doesn't settle its function in humans by any means, but does suggest numerous possibilities that have good liklihood of proving out experimentally. I have proposed an application of this to the human prion gene that is designed to yield reactive substrate analogues of potential use in the treatment of CJD.

So it behooves us to consider which phylogenetic intermediates between human and yeast are appropriate. DNA libraries from these bridge organims should be readily available to researchers. The list should generally avoid inbred domesticated stock because these may contain artifacts. It is important to remember that there are no 'lower' or 'primitive' organisms -- all extant organisms have been evolving for the same amount of time. None the less, organisms branching off earlier may retain important clues about the state of the gene at the time of divergence with the lineage that lead to mammals. Reconstructions methods can be validated directly to a certain extent by sequencing DNA from fossils of various types and ages.

An important by-product of sequencing the gene is information on how domain structure/function changed over time. Some models of evolutionary change envision building up protein function by shuffling in domains from other proteins. Prion genes have been extraordinarily conserved in mammals -- is it associated with advances or expansions in the central nervous system? Myelinated neurons arose 440 million years ago -- was the prion gene already 'mature' at this time? Or had the sequence settled down already in the notochords?

As an example, consider the situation with prion protein, a human gene of unknown function. The gene has been sequenced from perhaps 30 primates, 10 artiodactyls, 5 other vertebrates, 1 marsupial, and 1 bird. This list has failed to generate a probe that can identify a homologue iln nematode, fruit fly or yeast -- the gap is too big. In hindsight, far fewer species, better selected, might have linked this gene up to its superfamily years ago and we might now its normal role in humans by now.

The idea now is develop a list of species that efficiently remove reconstructive ambiguity from existing early nodes and provide the required access to the superfamily of this presently isolated gene. Weak nodes are quantitated and organisms to sequence are prioritized. A dynamic interactive process recomputes needed sequences as new ones come in, so that only a minimal bridging set is eventually sequenced.

The current situation allows reconstruction of the metatherian-eutherian node with some reliability but the mammalian-herptile node only with much uncertainty. The octapeptide repeat of placental mammals is a nonapeptide in marsupials and a higher order hexapeptide repeat in birds -- what should the probe for nemotode be here? We are left short.

I propose that the next round of sequencing be done initially so as to gradually extend our knowledge of prion gene back as far as fish. Before fish can be done, reptiles, more birds, and amphibians need be considered.

The fish lines that need consideration are respectively coelacanth and lungfish, modern bony fish, primitive bony fish, and cartilaginous fish. Suggested representatives of these lines in common experimental use or in evolutionary studies are given below. The temporal order of sequencing shuld be the order of divergence from the terrestrial vertebrate line.

Eukaryota; Metazoa; Chordata; Vertebrata; Pisces; Gnathostomata;


Protopterus aethiopicus (marbled lungfish) osteichthyes; dipneusti

Polypterus sp (bichir) osteichthyes; sarcopterygii; polypteriformes

Latimeria chalumnae (coelacanth) osteichthyes; crossopterygii

Carassius auratus (goldfish) osteichthyes; actinopterygii; cypriniformes

Cyprinus carpio (common carp) osteichthyes; actinopterygii; cypriniformes

Brachydanio rerio (zebrafish) osteichthyes; actinopterygii; cypriniformes

Oreochromis aureus (chichlid) osteichthyes; actinopterygii; perciformes

Anguilla japonica (japanese eel) osteichthyes; actinopterygii; anguilliformes

Oncorhynchus keta (chum salmon) osteichthyes; actinopterygii; salmoniformes

Myoxocephalus scorpius (shorthorn sculpin) osteichthyes; actinopterygii; scorpaeniformes

Lepisosteus oculatus (spotted gar) osteichthyes; actinopterygii; semionotiformes

Amia calva (bowfin) osteichthyes; actinopterygii; amiiformes

Squalus sucklii (puget sound dogfish) chondrichthyes

Raja clavata (thornback ray) chondrichthyes

Heterodontus francisci (horn shark) chondrichthyes

Crossopterygii (lobe-finned fishes): coelacanth and  lungfishes
... Protopterus annectens, African lungfishes] sarcopterygian fish (Sarcopterygii)
... Neoceratodus forsteri, Australian lungfish] dipnoan fish (Dipnoi)

... Latimeria chalumnae, coelacanth] Actinistia, Crossopterygii

Actinopterygii (ray-finned fish) actinopterygian, 
    modern bony fish teleosts (eel, carp,flounder)
Polypterus is the sister-group of actinopterygians
Teleostean fish
... American eel (Anguilla rostrata)
... Salmo salar (salmon)
... Oncorhynchus mykiss (rainbow trout)
... Gasterosteus aculeatus
... Anguilla anguilla
... Fundulus heteroclitus
... Sebastolobus altivelis
... Seriola quinquiradiata).marine teleost
... Cyprinus carpio teleost freshwater carp goldfish family. 
... Carassius auratus goldfish (Cypriniformes) ostariophysan tetraploidization 15-20 Mya
... Carassius carassius carp, cyprinid fishes
... Redanio zebrafish

Neoteleost acanthopterygian orders
Perciformes 
... Hexagrammos stelleri (Perciformes)
... Ernogrammus hexagrammus (Perciformes)
... Oreochromis spilurus (Perciformes) freshwater cichlid 
Scorpaeniformes
... Myoxocephalus stelleri (Scorpaeniformes)

Holostean fish
 ...Amia calva (Halecomorphi)bowfin  primitive bony fish, sister within the teleosts,
... Lepisosteus platyrhincus holostean fish 
Neoptergii. sturgeon is a sister to Neopterygii

Chondrichthytes
	Squalomorph sharks: superorder of elasmobranchs
	... Squalus acanthias shark 
	Batoidea rays and skates: superorder of elasmobranchs
	... Raja erinacia 
	... Raja tengu
	... Raja rhina skate

	Galeomorph sharks:
	... Heterodontus francisci
	... Carcharhinus obscurus



Siberian sturgeon (Acipenser baeri) (representative of a branch between
elasmobranchs and teleosts
(Cypriniformes) ostariophysan teleost tetraploidization 15-20 Mya
gene duplication by tetraploidy in teleost fish provide evidence that the mammalian ancester underwent a round
or two of tetraploid evolution, presumably at the stage of fish. It is possible that we might soon be able to deduce
the genomic structure of the Devonian fish ancestor. 
sea lamprey, Petromyzon marinus (agnathan)
....Rhipidistia extinct coelacanth closest relatives of tetrapods
Haemoglobins of bony fish match larval than adult amphibians