Cloning and characterization of ApRab4, a recycling Rab protein of Aiptasia pulchella, and its implication in the symbiosome biogenesis.
The biogenesis of Symbiodinium symbiosome in the host cells of the sea anemone, Aiptasia pulchella, involves retention of ApRab5 on and exclusion of ApRab11 from the organelle. One predicted consequence of this differential Rab association is the constant membrane fusion of symbiosomes with endocytic vesicles in the absence of parallel membrane retrieval and the subsequent formation of spacious symbiosomes, which nevertheless, contradicts the common perception. To solve this discrepancy, we determined whether membrane fusion occurs between symbiosomes and endocytic vesicles, and whether ApRab11-independent recycling is involved in symbiosome biogenesis. By using the biotin-avidin detection system, we found evidence for symbiosome-endocytic vesicle fusion. Cloning and characterization of ApRab4, an A. pulchella homolog of Rab4, showed that ApRab4 is associated with both the early endocytic and the perinuclear recycling compartments, and its normal function is required for the organization of the recycling compartments. Immunostaining localized ApRab4 to the symbiosome membrane, partially overlapping with ApRab5-decorated microdomains. Significantly, a treatment that impaired Symbiodinium photosynthesis also abolished symbiosome association of ApRab4. Furthermore, ApRab4 was quickly recruited to newly formed phagosomes, but prolonged association only occurred in those harboring live zooxanthelllae. We propose that ApRab4 retention on the symbiosome is an essential part of the mechanism for the biogenesis of Symbiodinium symbiosome.
Hong
Ming-Cheng
MC
Institute of Marine Biology, National Sun Yat-sen University, Kaohsiung, 804, Taiwan, Republic of China.
Huang
Yung-Sen
YS
Song
Pa-Ching
PC
Lin
Wen-Wen
WW
Fang
Lee-Shing
LS
Chen
Ming-Chyuan
MC
D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D005455:Fluorescent Antibody Technique; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D012615:Sea Anemones / Q000235:genetics*; D017422:Sequence Analysis, DNA; D013559:Symbiosis; D022161:Transport Vesicles / Q000378:metabolism; D014617:Vacuoles / Q000737:chemistry* / Q000378:metabolism; D020695:rab4 GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism
D017931:DNA Primers; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D020695:rab4 GTP-Binding Proteins