Genetic, genomic, and functional analysis of the granule lattice proteins in Tetrahymena secretory granules.
In some cells, the polypeptides stored in dense core secretory granules condense as ordered arrays. In ciliates such as Tetrahymena thermophila, the resulting crystals function as projectiles, expanding upon exocytosis. Isolation of granule contents previously defined five Granule lattice (Grl) proteins as abundant core constituents, whereas a functional screen identified a sixth family member. We have now expanded this screen to identify the nonredundant components required for projectile assembly. The results, further supported by gene disruption experiments, indicate that six Grl proteins define the core structure. Both in vivo and in vitro data indicate that core assembly begins in the endoplasmic reticulum with formation of specific hetero-oligomeric Grl proprotein complexes. Four additional GRL-like genes were found in the T. thermophila genome. Grl2p and Grl6p are targeted to granules, but the transcripts are present at low levels and neither is essential for core assembly. The DeltaGRL6 cells nonetheless showed a subtle change in granule morphology and a marked reduction in granule accumulation. Epistasis analysis suggests this results from accelerated loss of DeltaGRL6 granules, rather than from decreased synthesis. Our results not only provide insight into the organization of Grl-based granule cores but also imply that the functions of Grl proteins extend beyond core assembly.
Cowan
Andrew T
AT
Department of Molecular Genetics and Cell Biology, The University of Chicago, Chicago, IL 60637, USA.
Bowman
Grant R
GR
Edwards
Kyle F
KF
Emerson
J J
JJ
Turkewitz
Aaron P
AP
D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000235:genetics* / Q000502:physiology; D005089:Exocytosis / Q000502:physiology; D018503:Genome, Protozoan; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000378:metabolism; D017433:Protein Structure, Secondary; D015800:Protozoan Proteins / Q000235:genetics* / Q000502:physiology; D012270:Ribosomes / Q000502:physiology; D022142:Secretory Vesicles / Q000235:genetics* / Q000502:physiology; D016808:Tetrahymena thermophila / Q000235:genetics* / Q000502:physiology
D002135:Calcium-Binding Proteins; C103333:GRL1 protein, Tetrahymena thermophila; D011498:Protein Precursors; D015800:Protozoan Proteins