AN INITIATIVE TO SEQUENCE THE GENOME OF THE FILAMENTOUS FUNGUS ASPERGILLUSNIDULANS.
SUMMARYOn August 18 and 19th, an international (US, Europe and Japan) group offungal researchers met at the Fungal Genome Workshop in Stillwater Oklahomato consider the rationale and possible strategies for large scale genomicstudies of a filamentous fungus. Also present at this meeting were severalrepresentatives from industry as well as representatives from the yeast andhuman genome sequencing programs. This document presents a summary of thefindings of this meeting by an appointed Steering Committee of academic andindustrial scientists. The central findings of the committee can besummarized in three major points.
The fungal researchers present at Stillwater, reached a consensus that atwo stage strategy for completing the genomic sequence of Aspergillusnidulans be undertaken. In the first stage of the p!
roject, industrialsupport will be sought to fund the sequ!
encing of a large number ofexpressed sequence tags (ESTs) and the complete sequence of chromosome IV,the smallest A. nidulans chromosome, by a genome sequencing center. Duringthis stage, an international consortium of research laboratories inpartnership with established genome sequencing centers will prepare aconsortium grant for completing the A. nidulans genome sequence in fouryears.
- The properties of filamentous fungi as model genetic organisms;important producers of industrial enzymes, specialty chemi!
cals, andpharmaceuticals and their significant impact on human welfare as agents ofbiodegradation, spoilage and decay; provide a compelling rationale forlarge scale genome research. The information from the whole genomesequence would dramatically improve the utility of filamentous fungi inacademic and industrial research.
- The recent discoveries of several important genes and gene complexestogether with the availability of a complete physical map strongly arguesthat Aspergillus nidulans be chosen as the "composite" filamentous fungusfor large scale genomic study.
- In considering various strategies the group reached a consensus thatall efforts be directed towards obtaining the complete genome sequence inas short a time as possible.
WHY SEQUENCE A GENOME ?Genetic exploration of an organism traditionally relied on decades ofresearch and generations of researchers and geneticists. High through-putDNA sequencing now offers a rapid and cost effective approach to obtainingthe most important and relevant of all genetic information -- the completeDNA sequence of an organism. This technology has revolutionized biologicalresearch by moving science from a gene oriented science (how does this genework and what does this gene do ?) to a genome oriented scienc!
e (whatfunctions are encoded by this organism, and what is it capable of ?).This technology is dramatically altering the biotechnology industry byplacing the treasure of evolutionary genetic history in a computerdatabase.
WHY FILAMENTOUS FUNGI ?Whole genome sequencing of several eubacteria, archea and yeast haveprovided a logistical framework for studying microorganisms with larger andmore complex genomes. Filamentous fungi represent the next logical stepfor whole genome sequencing efforts. Here we summarize the findingspresented at the Stillwater meeting which provide a compelling rationalefor obtaining the complete genome sequence of a filamentous fungus.
- Filamentous fungi have a genome size two to three times that of yeast.Although some of this size can be accounted for by larger centromeres and alower gene density per nucleotide length, it is also clear that filamentousfungi have more genes.
n a recent cDNA sequencing study of the filamentous fungus !
Magnaporthegrisea, 30% of the genes contained no match in the current data bases anddid not match sequences in the yeast genome (B. Valent, Dupont Company).
- Several filamentous fungi are important genetic systems for the study ofgene regulation, cell cycle progression, recombination and development.
- Filamentous fungi are the most damaging crop pathogens and severalspecies cause life-threatening infections in humans.
- The metabolic versatility of filamentous fungi as compared to yeast iswell documented. For example, Aspergillus nidulans can utilize extremelydiverse nitrogen and carbon sources such as formamide and quinic acid.Growth can occur over broad ranges of temperature (10 to 45 C); pH (2.5 to11) and salt concentrations (up to 4.0M NaCl).
- Numerous industrial processes utilize enzymes and specific metabolitesproduced by filamentous fungi.These findings led to a consensus among academic and industrial researchers!
that the complete genome sequence of a filamentous fungus should beobtained.
WHY Aspergillus nidulans ? Aspergillus nidulans and Neurospora crassa are the most widely usedfilamentous fungi for genetic research. Uniquely, A. nidulans is a memberof a large and diverse genus whose members include some of the mostimportant filamentous fungi. Unlike Aspergilli A. nidulans has asophisticated sexual genetic system. Recent findings suggest its genomeencodes many of the activities of its industrially and medically importantrelatives.
Recent research findings.
For these reasons we propose that A. nidulans represents the best compositeorganism for fungal genome research.
- Like A. flavus or A. parasiticus, A. nidulans contains the entire aflatoxinbiosynthetic cluster (minus the last enzymatic step). Research shows thatA. nidulans can be used as a model system to study the regulation of thisgene cluster (T. Adams, N. Keller, Texas A&M Univers!
- Like Penicillin sp., A. nidulans!
contains the entire penicillinbiosynthetic cluster and can be genetically engineered for high levelproduction of penicillin. (G. Turner, University of Sheffield, England).
- Like A. fumigatus, A. nidulans can be used a model for invasive pulmonaryaspergillosis in neutropenic mice (D. Holden, Royal Postgraduate MedicalCollege, London).
- Transformation and genetic technologies developed in A. nidulans have beensuccessfully applied to a number of filamentous fungi for the purposes ofrecombinant enzyme production (Genencor Int., Novonordisk Biotech).
- A physical map of the A. nidulans genome has recently been completed. Thismap provides a sequence-ready road map for directed sequencing efforts (J.Arnold, R. Prade, University of Georgia).
- Finally, the organizational effort for the physical mapping project hasprepared the A. nidulans research community to take on the larger scaleproject of whole genome !
sequencing. The Aspergillus research community hasthe expertise in place to assume this task and is poised to use thisinformation to its best adavantage.
WHO WILL BENEFIT FROM THIS RESEARCH ?Academia and industry will both benefit.
Currently 25% of the known human disease genes have a homolog in yeast. Thecomplete genome sequence of A. nidulans will provide another powerfulgenetic system in which to study human metabolic disorders. The recent useof A. nidulans genetics to discover the human gene for Garrod's alkatonuria(M. Penalva, CSIC, Madrid) underscores this point.
The specialty chemical market is estimated at $15 billion annually. Thecomplete genome sequence of A. nidulans would facilitate cost effectivemetabolic engineering in industrially important ascomyce!
tes (N.Dunn-Coleman, Genencor International).
mparison to yeast or Drosophila, the number of researchers working onall filamentous fungi is small -- despite their obvious importance Forexample approximately 600 researchers are found in the Fungal Genetic StockCenter address list for each year from 1993 to 1996. In the U.S.,competition for funding has reduced the number of laboratories funded bythe National Institutes of Health and/or the National Science Foundation.A complete genome sequence would strengthen the filamentous fungalcommunity and significantly improve the ability of academic researchers tocompete for funding. More importantly, this research would help to sustainan active research community and potentially draw more researchers to thefield.
CONCLUSIONS.The Stillwater meeting provided a strong and cohesive opinion that the timeto begin a systematic approach to sequencing a fungal genome is NOW. Theparticipants were united in the opinion th!
at the genome of Aspergillusnidulans should be sequenced first.
There was a unanimous agreement among the participants that the trulyunique genetic and industrial importance of filamentous fungi should fosteran academic and industrial partnership to complete the task.
Next, we describes a two part strategy to achieve this end. We arrived atthis strategy after open discussions at the Stillwater meeting and aftercareful consultation with experienced genome researchers.
A TWO PART STRATEGY FOR COMPLETING THE GENOME SEQUENCE OF Aspergillus nidulans.PART 1: Establishing an expressed sequenced tag (EST) database andobtaining the complete sequence of chromosome IV.
We are currently seeking Industrial Partners (IPs) to fund the sequencingof a large number of EST's and the complete sequencing of chromosome IV.Dr. Bruce Roe (University of Oklahoma, Norman) in collaboration with Dr.Rolf Prade (Oklah!
oma State University) have agreed to perform this work at!
a cost of approximately $500,000. The time to completion is estimated atone year. EST's would provide an immediate gene resource and the sequenceof the smallest chromosome would provide a pilot project for completing theentire sequence. Industrial support is sought so that sequencing can beginin earnest. We are requesting the IPs consider supporting the project atthe level of one postdoctoral equivalent. Any additional support will beused the expand the initial sequencing effort and coordinate Part II(described below) by sponsoring a second Fungal Genome Workshop.
We are asking that IPs indicate their willingness to support this projectby writing to:
Steering Committee for the Fungal Genome ProjectC/O Dr. Gregory May
Dept. of Cell Biology
Baylor College of Medicine
One Baylor Plaza
Houston, TX 77030
PHONE: (713) 798-4756
FAX: (713) 798-7799
Upon the identification!
of potential IPs, Drs. Prade and Roe will be askedto prepare a detailed budget, and a concise research plan including adescription of how the sequence data will be disseminated. The SteeringCommittee will facilitate negotiations between the IP's and Drs. Prade andRoe. Research progress will be gauged in 6 months at a second FungalGenome Workshop. This Workshop will also be used to organize Part II ofthe project.
PART II. Establishing a consortium for completing the A. nidulans genomesequence.
The sequence of chromosome IV will provide preliminary data for a moresubstantial proposal to complete the genome sequence. Over the next yearthe Steering Committee will:
- Identify fungal researchers from the US, Europe and Japan who arewilling to participate in this project. It is important that the fungalcommunity be directly involved in sequencing efforts rather than justwaiting to be fed the data. Participation by f!
ungal research laboratorieswill foster the development of!
expertise which is expected to feedadditional fungal sequencing efforts. Each fungal group involved willestablish a collaboration with a genome sequencing center to facilitaterapid technology development. Encouraging support from several centers andgenome based companies has already been obtained (Bruce Roe, TIGR, SangerCenter, Genome Therapeutics). If necessary the Steering Committee willfacilitate contacts between academic groups and laboratories withlarge-scale genome sequencing capability.
- Coordinate efforts between sequencing groups. Specific experimentalapproaches (e.g. shotgun cloning of chromosomes, deep-end BAC sequencing,directed sequencing) will be left to the discretion of the research groupsand their collaborators. The Steering Committee will ensure that theentire genome will be covered.
- Identify researchers who are willing to serve as informatics managersfor the project. The Steering Committee recognizes that making !
thesequence available to the research community in a useable format
willrequire considerable informatics support.
funding sources (Public and Private) for the entire project.The
estimate cost based on $0.50 per nucleotide and a 30 Mb genome is
- Greg May, Baylor College of
- Tom Adams, Texas A&M
- Jonathan Arnold, University of
- Claudio Scazzocchio, University of Paris
Turner, University of Sheffield
- Jeff Shuster, NovoNordisk
- Nigel Dunn-Coleman, Genencor International
- John E. Hamer, Purdue University