%A Liu,Shijie %A Majeed,Waqar %A Kudlyk,Tetyana %A Lupashin,Vladimir %A Storrie,Brian %D 2016 %J Frontiers in Cell and Developmental Biology %C %F %G English %K Golgi organization,Rab41/6d,effector,dynactin 6,Syntaxin 8,Rab6a/a’ %Q %R 10.3389/fcell.2016.00013 %W %L %M %P %7 %8 2016-March-01 %9 Original Research %+ Brian Storrie,Department of Physiology and Biophysics, University of Arkansas for Medical Sciences,Little Rock, AR, USA,storriebrian@uams.edu %# %! Rab41/6d effectors and Golgi organization %* %< %T Identification of Rab41/6d Effectors Provides an Explanation for the Differential Effects of Rab41/6d and Rab6a/a' on Golgi Organization %U https://www.frontiersin.org/articles/10.3389/fcell.2016.00013 %V 4 %0 JOURNAL ARTICLE %@ 2296-634X %X Unexpectedly, members of the Rab VI subfamily exhibit considerable variation in their effects on Golgi organization and trafficking. By fluorescence microscopy, neither depletion nor overexpression of the GDP-locked form of Rab6a/a', the first trans Golgi-associated Rab protein discovered, affects Golgi ribbon organization while, on the other hand, both Rab41/6d depletion and overexpression of GDP-locked form cause Golgi fragmentation into a cluster of punctate elements, suggesting that Rab41/6d has an active role in maintenance of Golgi ribbon organization. To establish a molecular basis for these differences, we screened for Rab41/6d interacting proteins by yeast two-hybrid assay. 155 non-repetitive hits were isolated and sequenced, and after searching in NCBI database, 102 different proteins and protein fragments were identified. None of these hits overlapped with any published Rab6a/a' effector. Eight putative Rab41 interactors involved in membrane trafficking were found. Significantly, these exhibited a preferential interaction with GTP- vs. GDP-locked Rab41/6d. Of the 8 hits, the dynactin 6, syntaxin 8, and Kif18A plasmids were the only ones expressing the full-length protein. Hence, these 3 proteins were selected for further study. We found that depletion of dynactin 6 or syntaxin 8, but not Kif18A, resulted in a fragmented Golgi apparatus that displayed a Rab41/6d knockdown phenotype, i.e., the Golgi apparatus was disrupted into a cluster of punctate Golgi elements. Co-immunoprecipation experiments verified that the interaction of dynactin 6 and syntaxin 8 with GTP-locked Rab41/6d was stronger than that with wild type Rab41/6d and least with the GDP-locked form. In contrast, co-immunoprecipitation interaction with Rab6a was greatest with the GDP-locked Rab6a, suggestive of a non-physiological interaction. In conclusion, we suggest that dynactin 6, a subunit of dynactin complex, the minus-end-directed, dynein motor, provides a sufficient molecular basis to explain the active role of Rab41/6d in maintaining Golgi ribbon organization while syntaxin 8 contributes more indirectly to Golgi positioning.