NblA, a Key Protein of Phycobilisome Degradation, Interacts with ClpC, a HSP100 Chaperone Partner of a Cyanobacterial Clp Protease

Abstract

When cyanobacteria are starved for nitrogen, expression of the NblA protein increases and thereby induces proteolytic degradation of phycobilisomes, light-harvesting complexes of pigmented proteins. Phycobilisome degradation leads to a color change of the cells from blue-green to yellow-green, referred to as bleaching or chlorosis. As reported previously, NblA binds via a conserved region at its C terminus to the ␣-subunits of phycobiliproteins, the main components of phycobilisomes. We demonstrate here that a highly conserved stretch of amino acids in the N-terminal helix of NblA is essential for protein function in vivo. Affinity purification of glutathione S-transferase-tagged NblA, expressed in a Nostoc sp. PCC7120 mutant lacking wildtype NblA, resulted in co-precipitation of ClpC, encoded by open reading frame alr2999 of the Nostoc chromosome. ClpC is a HSP100 chaperone partner of the Clp protease. ATP-dependent binding of NblA to ClpC was corroborated by in vitro pulldown assays. Introducing amino acid exchanges, we verified that the conserved N-terminal motif of NblA mediates the interaction with ClpC. Further results indicate that NblA binds phycobiliprotein subunits and ClpC simultaneously, thus bringing the proteins into close proximity. Altogether these results suggest that NblA may act as an adaptor protein that guides a ClpC⅐ClpP complex to the phycobiliprotein disks in the rods of phycobilisomes, thereby initiating the degradation process.

Degradation of Phycobilisomes in Synechocystis sp. PCC6803

“…Binding experiments have shown that NblA interacts with the ␣-subunits of phycobiliproteins (18,19). In pull-down experiments, NblA was found to bind to ClpC, an HSP100 chaperone partner of a Clp protease, in an ATP-dependent manner (20). This result led to a proposed model of PBS degradation in which NblA acts as a so-called adaptor protein of a Clp protease (20).…”

“…In pull-down experiments, NblA was found to bind to ClpC, an HSP100 chaperone partner of a Clp protease, in an ATP-dependent manner (20). This result led to a proposed model of PBS degradation in which NblA acts as a so-called adaptor protein of a Clp protease (20). Clp degradation complexes consist of two functional elements: a cylinder-like proteolytic core of two heptameric rings and an AAAϩ chaperone (21,22).…”

“…Sequence alignment of NblA proteins reveals a low homology (about 30% sequence identity on average) (27). Short stretches of highly conserved amino acid residues located near the N and C termini of the proteins are involved in phycobiliprotein and ClpC binding (19,20). NblA proteins from Nostoc sp.…”

Nitric Oxide–Triggered Remodeling of Chloroplast Bioenergetics and Thylakoid Proteins upon Nitrogen Starvation inChlamydomonas reinhardtii

“…Last, the signaling pathway may include some adaptor proteins of proteases that recognize and bind both the substrates for degradation and the chaperone subunit of the protease (reviewed in Kirstein et al, 2009). For instance, in cyanobacteria, the NblA adaptor mediates the proteolytic degradation of phycobilisomes during nitrogen or sulfur starvation because of its double affinity for phycobilisome rods and for ClpC (Collier and Grossman, 1994;Karradt et al, 2008).…”

Degradation of SsrA-tagged proteins in streptococci

“…Gram-positive bacteria such as S. aureus or B. subtilis do not encode homologues of SspB, although they contain ClpX. However, several adaptor proteins for ClpC have SsrA-tagged proteins in streptococci been identified in Gram-positive bacteria, including MecA, McsB, NblA, YpbH and TrfA (Andersson et al, 2006;Donegan et al, 2014;Karradt et al, 2008;Kirstein et al, 2007;Persuh et al, 2002;Turgay et al, 1998). All known ClpC activities require the participation of an adaptor protein (Kirstein et al, 2007(Kirstein et al, , 2009Schlothauer et al, 2003;Turgay et al, 1998;Wang et al, 2011).…”