madrid2i

Resumen

The present invention relates to an isolated nucleic acid sequence coding for a PntR transcription factor comprising a nucleotide sequence that is the same as, or is complementary to, or contains any suitable codon substitutions for any of those of SEQ ID NOs: 1, 3 or 5 or comprises a sequence which has at least 60% sequence homology with any of SEQ ID NOs: 1, 3 or 5, as well as corresponding polypeptides, vector systems and host cells comprising the same. In particular, the invention relates to sequences which are obtained from an Aspergillus, Trichoderma or Penicillium cell. The invention further relates to methods for disrupting PntR expression in a cell as well as methods for expression or production of a POI in a host cell in which PntR expression has been disrupted.

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CLAIMS 1. An isolated nucleic acid sequence coding for a PntR transcription factor comprising a nucleotide sequence that is the same as, or is complementary to, or contains any suitable codon substitutions for any of those of SEQ ID NOs: 1, 3 or 5 or comprises a sequence which has at least 60% sequence homology with any of SEQ ID NOs: 1, 3 or 5. 2. An isolated nucleic acid sequence according to claim 1 which encodes a PntR transcription factor polypeptide comprising the amino acid sequence as shown in any of SEQ ID NOs: 2, 4 or 6 or a sequence having at least 75% identity thereto or an effective fragment thereof. 3. An isolated nucleic acid sequence according to claim 2 which is obtained from an Aspergillus, Trichoderma or Penicillium cell. 4. An isolated nucleic acid sequence according to claim 3 which is obtained from an Aspergillus* cell. 5. A plasmid or expression vector system comprising a nucleic acid sequence according to any of claims 1 to 4. 6. A host cell transformed or transfected with the plasmid or expression vector system of claim 5. 7. A polypeptide comprising the amino acid sequence corresponding to Aspergillus PntR or a functional equivalent thereof. 8. A polypeptide according to claim 7 having the same amino acid sequence or a sequence that is at least 75% identical to that of an Aspergillus PntR. 9. A polypeptide according to claim 7 or claim 8 comprising the amino acid sequence as shown in any of SEQ ID NOs: 2, 4 or 6 or a sequence having at least 75% identity thereto or an effective fragment thereof. 10. A nucleic acid construct arranged to knock down the expression or function of the transcription factor PntR. 11. A nucleic acid construct according to claim 10 comprising a nucleic acid sequence encoding a functionally disrupted PntR. 12. A nucleic acid construct according to claim 11 comprising a nucleic acid sequence encoding PntR and lacking the region encoding Zn2Cy6. 13. A nucleic acid construct according to claim 10 comprising all or a fragment of a nucleic acid sequence of any of claims 1 to 4. 14. A plasmid or vector system comprising a nucleic acid construct according to any of claims 10 to 13. 15. A method of disrupting PntR expression in a cell comprising introducing a nucleic acid construct as claimed in any of claims 10 to 13 or a plasmid or vector system as claimed in claim 14 into said cell. 16. A host cell transformed or transfected with a nucleic acid construct arranged to knock down the expression or function of the transcription factor PntR as claimed in any of claims 10 to 13 or a plasmid or vector system according to claim 14. 17. A host cell according to claim 16 further comprising a nucleic acid construct comprising a coding sequence of a protein of interest (POI) under the transcriptional control of a promoter regulated by XInR. 18. A host cell according to claim 16 or claim 17 wherein said host cell is selected from the group consisting of Aspergillus sp., Trichoderma and Penicillium sp. 19. A host cell according to any of claims 16 to 18 capable of producing an extracellular xylanase. 20. A host cell according to any of claims 16 to 19 wherein said cell is protease-deficient. 21. A host cell according to any of claims 16 to 20 wherein said cell has a block in L-arabinose metabolism. 22. A host cell according to any of claims 16 to 21 wherein said cell exhibits increased xylanase activity. 23. A host cell according to any of claims 16 to 22 wherein said cell is useful for production of a POI. 24. A method of expressing in a host cell a POI whose coding sequence is under the transcriptional control of a promoter regulated by XInR, said method comprising modulating the transcription factor PntR in said host cell. 25. A method according to claim 24 comprising the steps of providing a PntR-deficient host cell comprising the gene encoding the POI under XInR transcriptional control, and cultivating the host cell under suitable conditions. 26. A method according to any of claims 24 to 25 wherein the PntR has a nucleic acid sequence as set out in any of SEQ ID NOS: 1 , 3 or 5 or a sequence having more than 60% identity thereto. 27. A method according to any of claims 24 and 26 comprising using a host cell according to any of claims 16 to 23. 28. A method according to any of claims 24 to 27 wherein the POI is a homologous protein. 29. A method according to any of claims 24 to 27 wherein the POI is a heterologous protein. 30. A method according to any of claims 24 to 29 wherein the host cell is a filamentous fungal cell and, preferably, Aspergillus, Trichoderma or Penicillium spp. 31. A method according to any of claims 24 to 30 wherein said method of expressing a POI is part of a method of production of said POI and further comprisesisolating/purifying said POI. 32. A method of production of a POI comprising expressing in a host cell a POI whose coding sequence is under the transcriptional control of a promoter regulated by XInR, said method comprising modulating the transcription factor PntR in said host cell. 33. Use of modulation of the transcription factor PntR in the manufacture of a host cell to increase expression of a protein of interest (POI) whose coding sequence is under the transcriptional control of a promoter regulated by XInR.

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