Characterization of Hap/BAG-1 variants as RP1 binding proteins with antiapoptotic activity.
The MAPRE protein family (EB1, RP1, EB2) represents a highly conserved group of proteins that localize preferentially to the plus end of microtubules, both in the nucleus and cytoplasm. In addition, MAPRE family members are characterized by their capability to bind to the C-terminus of the adenomatous polyposis coli (APC) protein and tubulin in order to stabilize microtubules. Apart from the interaction with APC and tubulin, no other direct binding partners are known today. Because the RP1 gene product was identified in activated T cells, we set out to search for new interacting molecules in a yeast 2-hybrid system. We isolated a cDNA variant encoding for the antiapoptotic Hap/BAG-1 protein truncated by 34 amino acids at the C-terminus. In the original Hap/BAG-1 protein, the C-terminal domain is responsible for binding to Bcl-2 and Hsp/Hsc70, which is believed to be the reason for its antiapoptotic activity. Although this putative Hap/BAG-1 variant protein showed no interaction with Bcl-2 or Hsp/Hsc70, it was perfectly able to confer resistance to apoptosis. Subcellular distribution analysis revealed that the Hap/Bag-1 variant protein localized homogenously to the cytoplasm and shuttles into the nucleus in response to stress, a process that could be blocked by RP1 protein overexpression.
Wadle
Andreas
A
Med. Department I, Saarland University Medical School, Homburg, Germany.
Mischo
Axel
A
Henrich
Philipp P
PP
Stenner-Liewen
Frank
F
Stenner-Lieven
Frank
F
Scherer
Christoph
C
Imig
Jochen
J
Petersen
Gabriele
G
Pfreundschuh
Michael
M
Renner
Christoph
C
D000595:Amino Acid Sequence; D017209:Apoptosis; D015153:Blotting, Western; D002352:Carrier Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D002460:Cell Line; D002467:Cell Nucleus / Q000737:chemistry; D003593:Cytoplasm / Q000737:chemistry; D018076:DNA, Complementary / Q000737:chemistry; D004268:DNA-Binding Proteins; D005136:Eye Proteins / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D005982:Glutathione Transferase / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D007163:Immunosorbent Techniques; D008869:Microtubule-Associated Proteins; D008969:Molecular Sequence Data; D009842:Oligopeptides; D010455:Peptides / Q000235:genetics; D011993:Recombinant Fusion Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors; D014162:Transfection; D020798:Two-Hybrid System Techniques; D015003:Yeasts / Q000235:genetics
C091732:BCL2-associated athanogene 1 protein; D002352:Carrier Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D005136:Eye Proteins; D008869:Microtubule-Associated Proteins; D009842:Oligopeptides; D010455:Peptides; C120042:RP1 protein, human; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; C085706:FLAG peptide; D005982:Glutathione Transferase