Addendum

We recently showed (Bonazzi et al. 7, 570–580; 2005) that the protein CtBP3/BARS (BARS) controls basolateral but not apical traffic from the Golgi to the plasma membrane. One of the several lines of evidence supporting this conclusion was that a 'smart pool' of siRNA duplexes (Dharmacon, Lafayette, CA) against the human sequence of BARS caused the selective knockdown of BARS and the inhibition of basolateral but not of apical traffic in COS-7 cells (Fig. 3e; Bonazzi et al.; 2005). Because the BARS nucleotide sequence of COS-7 cells (of monkey origin) is unknown, we have now: (1) obtained the COS-7 BARS sequence (V. Hsu, personal communication); and (2) performed rescue-of-function experiments in siRNA-treated COS-7 cells by restoring their BARS cytosolic levels with recombinant rat protein.

We report that: (1) COS-7 BARS contains a sequence (CCGTCAAGCAGATGAGACA) that perfectly matches one of the siRNA oligonucleotides in the Dharmacon pool, thus justifying the published effects of this pool in COS-7 cells; and (2) COS-7 cells that were depleted of BARS by siRNA and then were treated exactly as described in the experiment in Fig. 3e (Bonazzi et al.; 2005) regain normal levels of basolateral transport to the plasma membrane upon microinjection with recombinant BARS (Table 1). These results confirm the specificity of the siRNA effects on basolateral traffic shown in Fig. 3e (Bonazzi et al.; 2005).

Table 1 Specificity of the effects of siRNAs against BARS on VSVG traffic