Tubulo-vesicular structure localization signals

Abstract

The invention relates to polarized cell tubulo-vesicular structure localization signals. The localization signals are utilized as research tools or are linked to polypeptides of interest or therapeutic molecules. Disclosed are methods of making and using polypeptides and modified polypeptides as signals to localize therapeutics, experimental compounds, peptides, proteins and/or other macromolecules to the tubulo-vesicular structures of polarized cells. The polypeptides of the invention optionally include linkage to reporters, epitopes and/or other experimental or therapeutic molecules. The invention also encompasses polynucleotides encoding the localization signals and vectors comprising these polynucleotides.

1 . A polypeptide containing an amino acid sequence at least 80% identical to SEQ ID NO:5 in frame with an amino acid sequence at least 80% identical to SEQ ID NO:6. 
     
     
         2 . A polypeptide containing an amino acid sequence at least 80% identical to SEQ ID NO:7 in frame with an amino acid sequence at least 80% identical to SEQ ID NO:8. 
     
     
         3 . A combination localization signal comprising a peptide at least 80% identical to a subsequence of SEQ ID NO:1 in frame with a peptide at least 80% identical to a subsequence of subsequence of SEQ ID NO:2. 
     
     
         4 . A combination localization signal comprising a peptide at least 80% identical to a subsequence of SEQ ID NO:3 in frame with a peptide at least 80% identical to a subsequence of subsequence of SEQ ID NO:4. 
     
     
         5 . A tubulo-vesicular structure localization signal comprising at least one copy of a peptide selected from the group consisting of:
 a) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 1-145 of the amino acid sequence of SEQ ID NO:1;   b) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 1-140 of the amino acid sequence of SEQ ID NO:1;   c) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 1-135 of the amino acid sequence of SEQ ID NO:1; and   d) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 1-130 of the amino acid sequence of SEQ ID NO:1.   
     
     
         6 . A localization signal comprising at least one copy of a peptide selected from the group consisting of:
 a) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 594-627 of the amino acid sequence of SEQ ID NO:2;   b) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 591-627 of the amino acid sequence of SEQ ID NO:2;   c) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 584-627 of the amino acid sequence of SEQ ID NO:2; and   d) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 577-627 of the amino acid sequence of SEQ ID NO:2.   
     
     
         7 . A combination localization signal comprising a peptide of  claim 5  in frame with a peptide of  claim 6 . 
     
     
         8 . A localization signal comprising at least one copy of a peptide selected from the group consisting of:
 a) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 1-125 of the amino acid sequence of SEQ ID NO:3;   b) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 1-130 of the amino acid sequence of SEQ ID NO:3;   c) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 135 of the amino acid sequence of SEQ ID NO:3; and   d) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 1-140 of the amino acid sequence of SEQ ID NO:3.   
     
     
         9 . A localization signal comprising at least one copy of a peptide selected from the group consisting of:
 a) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 581-602 of the amino acid sequence of SEQ ID NO:4;   b) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 576-602 of the amino acid sequence of SEQ ID NO:4;   c) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 573-602 of the amino acid sequence of SEQ ID NO:4; and   d) a peptide at least 80% identical to a peptide comprising amino acid residues corresponding to amino acid residues 5769-602 of the amino acid sequence of SEQ ID NO:4.   
     
     
         10 . A combination localization signal comprising a peptide of  claim 8  in frame with a peptide of  claim 9 . 
     
     
         11 . The localization signal of  claim 3  linked to an epitope. 
     
     
         12 . The localization signal of  claim 3  linked to a reporter. 
     
     
         13 . The localization signal of  claim 3  linked to a polypeptide of interest. 
     
     
         14 . The localization signal of  claim 3  linked to a therapeutic molecule. 
     
     
         15 . A nucleic acid molecule comprising a polynucleotide sequence encoding at least one copy of the peptide of  claim 1 . 
     
     
         16 . A vector comprising the nucleic acid molecule of  claim 15 . 
     
     
         17 . A recombinant host cell comprising the vector of  claim 16 . 
     
     
         18 . A method of localizing a polypeptide of interest to a tubulo-vesicular structure subcellular compartment in a cell comprising linking a polypeptide of interest open reading frame to a nucleic acid molecule of  claim 15  to create a fusion protein coding sequence, and transfecting the fusion protein coding sequence into a host cell and culturing the transfected host cell under conditions suitable to produce at least one copy of the fusion protein.