Polynucleotides encoding patched proteins

• US 6,610,507 B2
• Filed: 10/20/1997
• Issued: 08/26/2003
• Est. Priority Date: 10/07/1994
• Status: Expired due to Fees

• Current Assignee: Board of Trustees of the Leland Stanford Junior University (Stanford Management Co.)
• Original Assignee: Board of Trustees of the Leland Stanford Junior University (Stanford Management Co.)
• Inventors: Johnson, Ronald L., Goodrich, Lisa V., Scott, Matthew P.
• Primary Examiner(s): Eyler, Yvonne
• Assistant Examiner(s): BRANNOCK, MICHAEL T
• Application Number:
  US08/954,701
• Publication Number:
  US 20030032085A1
• Time in Patent Office: 2,136 Days
• Field of Search: 435/6, 435/7.1, 435/69.1, 435/252.3, 435/320.1, 435/325, 435/7.21, 536/23.5, 536/24.1, 436/501, 530/350, 514/2
• US Class Current: 435/69.1
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/43563 from insects
• CPC Class Codes: C07K 14/47 from mammals
• CPC Class Codes: C07K 14/705 Receptors; Cell surface antigens; Cell surface determinants tumour specific antigens C07K14/4748

Abstract

Invertebrate and vertebrate patched genes are provided, including the mouse and human patched genes, as well as methods for isolation of related genes, where the genes may be of different species or in the same family. Having the ability to regulate the expression of the patched gene, allows for the elucidation of embryonic development, cellular regulation associated with signal transduction by the patched gene, the identification of agonist and antagonist to signal transduction, identification of ligands for binding to patched,isolation of the ligands, and assaying for levels of transcription and expression of the patched gene.

Claims

1. A nucleic acid including a nucleic acid sequence that hybridizes under stringent conditions, including a wash step of 0.2×

SSC at 65°

C., to a sequence selected from at least one of SEQ ID Nos. 9 or 18, wherein the nucleic acid sequence encodes an amino acid sequence that binds a naturally occurring hedgehog polypeptide, and wherein the nucleic acid sequence is not identical to SEQ ID No. 5 or a fragment thereof of at least 30 nucleotides.

2. A nucleic acid sequence according to claim 1, wherein the nucleic acid sequence is a mammalian nucleic acid sequence.

3. A nucleic acid sequence according to claim 2, wherein said nucleic acid sequence is a human sequence.

4. A nucleic acid sequence according to claim 2, wherein said nucleic acid sequence is a mouse sequence.

5. A nucleic acid sequence according to claim 1 joined to a nucleic acid sequence comprising a restriction enzyme recognition sequence.

6. An expression cassette comprising a transcriptional initiation region functional in an expression host, a nucleic acid sequence according to claim 1 under transcriptional regulation of said transcriptional initiation region, and a transcriptional termination region functional in said expression host.

7. An expression cassette according to claim 6, wherein said transcriptional initiation region is heterologous to the nucleic acid sequence.

8. An expression cassette according to claim 6, wherein said transcriptional initiation region is homologous to the nucleic acid sequence.

9. A cell comprising the expression cassette according to claim 6 as part of an extrachromosomal element or integrated into the genome of a host cell as a result of introduction of said expression cassette into said host cell and the cellular progeny of said host cell.

10. A cell according to claim 9, further comprising a polypeptide, provided by the expression cassette, in the cellular membrane of said cell.

11. A cell according to claim 10, wherein said polypeptide is a mouse polypeptide.

12. A cell according to claim 10, wherein said polypeptide is a human polypeptide.

13. A cell according to claim 9, wherein said transcriptional initiation region is a transcriptional initiation region endogenously associated with a Drosophila nucleic acid encoding SEQ ID No. 6 comprising a promoter and enhancer.

14. A cell comprising an expression cassette comprising a transcriptional initiation region functional in an expression host, a nucleic acid according to claim 1 under the transcriptional regulation of said transcriptional initiation region, a marker gene, and a transcriptional termination region functional in said expression host, wherein said transcriptional initiation region consists of a 5′

non-coding region regulating the transcription of said nucleic acid, as part of an extrachromosomal element or integrated into the genome of a host cell as a result of introduction of said expression cassette into said host, and cellular progeny thereof.

15. A cell according to claim 14, wherein said transcriptional initiation region is a transcriptional initiation region associated with a Drosophila nucleic acid encoding SEQ ID No. 6.

16. A method for producing a polypeptide comprising culturing a cell according to claim 9, whereby said cell expresses said polypeptide;

and purifying said polypeptide.

17. A nucleic acid including a nucleic acid sequence encoding an amino acid sequence that binds a naturally occurring hedgehog polypeptide, wherein the amino acid sequence is at least 90% identical to a sequence selected from at least one of SEQ ID No. 10 or 19.

18. A nucleic acid of claim 17, wherein the amino acid sequence is at least 95% identical to a sequence selected from SEQ ID No. 10 or 19.

19. A nucleic acid of claim 18, wherein the amino acid sequence is at least 98% identical to a sequence selected from SEQ ID No. 10 or 19.

20. A nucleic acid of claim 19, wherein the amino acid sequence is identical to a sequence selected from at least one of SEQ ID No. 10 or 19.

21. A nucleic acid sequence according to claim 17 joined to a nucleic acid sequence comprising a restriction enzyme recognition sequence.

22. An expression cassette comprising a transcriptional initiation region functional in an expression host, a nucleic acid sequence according to claim 17, under the transcriptional regulation of said transcriptional initiation region, and a transcriptional termination region functional in said expression host.

23. A cell comprising the expression cassette according to claim 22 as part of an extrachromosomal element or integrated into the genome of a host cell as a result of introduction of said expression cassette into said host cell and the cellular progeny of said host cell.

24. A method for producing a polypeptide comprising culturing a cell according to claim 23, whereby said cell expresses said polypeptide;

and purifying said polypeptide.