The biosynthesis of secretory proteins requires translocation through the Sec61 channel into the endoplasmic reticulum (ER) . The protein conducting channel is formed by three proteins and is conserved in evolution from archaea and bacteria to eukaryotic cells [2, 3]. The largest subunit forming the actual protein conducting channel is named Sec61α in eukaryotes and SecY in bacteria and archaea. The β-subunit is termed Sec61β in metazoans and SecG in bacteria and archaea, whereas in yeast Saccharomyces cerevisiae (S. cerevisiae) two homologous β-subunits, Sbh1p and Sbh2p exist. The γ-subunit is called Sec61γ in mammalian cells, Sss1p in S. cerevisiae cells and SecE in bacteria and archaea. In S. cerevisiae two homologous trimeric Sec61 complexes exist. The Sec61 complex is composed of Sec61p, Sbh1p and Sss1p [4–7]. This complex together with Sec63p and Kar2p functions in co-translational translocation . In association with Sec62p, Sec63p, Sec71p and Sec72p, Sec61 complex mediates also post-translational protein translocation as the so-called Sec complex [6, 9]. In addition to its role in co- and post-translational translocation, the Sec61 complex has also been implicated in retro-translocation of misfolded proteins to the cytosol for degradation [10, 11]. The second S. cerevisiae translocation complex, the Ssh1 complex, consists of the Sec61p homologue Ssh1p, the Sbh1p homologue Sbh2p, and Sss1p [2, 8]. The Ssh1 complex has been shown to contribute to both co- and post-translational translocation and ER associated degradation (ERAD) [4, 12].
Recent structural studies have shed light on the assembly and function of the α-subunit as the protein conducting channel [3, 13, 14]. In contrast, the functions of the β- and γ-subunits remain largely unclear. The β-subunit is not essential, but has a facilitating role in translocation in mammalian cells . In addition to its interactions with Sec61p, the mammalian β-subunit has been shown to interact with signal peptidase  and the ribosome . The yeast β-subunit, Sbh1p, was shown to act as a guanine nucleotide exchange factor for signal recognition particle receptor . The two yeast β-subunits, Sbh1p and Sbh2p are encoded by non-essential genes. Deletion of either gene alone has no effect on growth, while the deletion of both SBH1 and SBH2 results in temperature-sensitivity for growth [4, 7]. Different studies have reported variable effects of SBH1 and SBH2 deletion on protein translocation of the α mating pheromone precursor (ppαf), Kar2p, bacterial α-amylase and dipeptidyl aminopeptidase [4, 15, 18, 19]. These results indicate a role for Sbh1p in co-translational protein translocation. Interestingly, the Sbh1p tm-domain alone is sufficient to support in vivo co-translational protein translocation and for interactions with Sec61 both in S. cerevisiae and in S. pombe [18–20].
The non-essential role of SBH1 and SBH2, in contrast to the essential genes encoding the α- and γ-subunits Sec61p and Sss1p, suggests that the β-subunits may have functions that are not directly linked to protein translocation. Previously, a stabilizing role in the translocation complex has been suggested for both β- and γ-subunits . Both in mammalian and yeast cells, β-subunits have been shown to co-immunoprecipitate with the exocyst complex components Sec8p, Sec10p and Sec15p [22, 23] whose main function is thought to be tethering of transport vesicles to the plasma membrane . In addition, SBH1 over-expression has been shown to rescue the temperature-sensitive growth phenotype of several exocyst subunit mutations [22, 23].
It appears that a pool of Sbh1p exists outside Sec61 complex and that these Sbh1p proteins can interact with Rtn1p, a member of the reticulon family proteins . Rtn1p, together with Rtn2p and the DP1/Yop1p family protein Yop1p have been shown to function in modulation of the ER membrane structure [25–27]. Interestingly, in addition to its interactions with Sbh1p, Rtn1p has also been shown to co-purify with Exocyst subunit Sec6 . Collectively, the current data suggests that Sbh1p displays complex molecular interactions and that its function may not be restricted to protein translocation. In this study we characterize the molecular determinants that are important for the function of the Sec61 protein translocation complex β-subunit tm-domain function. In addition, we show that this β-subunit interacts with ER-resident reticulon protein complexes.