# Chloroplasts at the Crossroad of Photosynthesis, Pathogen Infection and Plant Defense
## 8 Chloroplasts are Targeted by Pathogen Effectors
### HopI1 and HopN1  
HopI1 and HopN1 are chloroplast-targeted _P. syringae_ T3 Es that enter the chloroplast via a noncanonical mechanism and are not processed after entry [306; 307; 320; 321; 328]. Confocal microscopic analysis of transiently expressed fluorescent protein-tagged effectors showed that HopI1 and HopN1 localize to the chloroplast [306; 307]. The chloroplast localization of these two T3 Es was further confirmed with subcellular fractionation [306; 307]. It should be noted that transiently expressed HopN1-YFP protein was previously found in the plasma membrane of Chinese cabbage and tobacco epidermal cells [341]. One possible reason for the discrepancy is the absence of chloroplasts in epidermal cells [307].  
HopI1 is a Dna] protein with a phosphate-binding loop (P-loop), a proline and glutamine (PQ)-rich repeat region, and a J-domain with a histidine-proline-aspartate (HPD) motif [306; 320], HopI1 was found to cause thylakoid membrane remodeling and suppression of SA accumulation and SA-dependent defenses [306]. As J-domains are known to interact with 70 kDa heat shock proteins (Hsp70s) and activate their ATPase activity and protein folding ability [342], Hsp70s were proposed to be the target of HopII [306]. Consistent with this hypothesis, the J-domain of HopI1 from _P. syringae_ pv. _maculicola_ strain ES4326 was found to bind to full-length plant cytHsp70-1 (cyt stands for cytosolic) in vitro, and full-length HopI1 successfully pulled down cpHSP70-1 (cp stands for chloroplast) from pea chloroplasts [321]. In addition, the J-domain of HopII stimulated the ATP hydrolysis activity of Hsp70s [321]. Furthermore, immunoprecipitation showed that HopI1 interacts with both cytHsp70-1 and cpHsp70-1, recruits cytHsp70-1 to the chloroplast, and forms large complexes with cytHsp70-1 and cpHsp70-1 [321]. The involvement of cytHsp70-1 in this process is interesting. It is possible that the level of cpHsp70-1 is not high enough for HopI1 to function without recruiting cytHsp70-1 to the chloroplast [321]. Indeed, genetics studies showed that cytHsp70-1 is required for the virulence role of HopII at standard growth temperatures [321]. Therefore, it was hypothesized that, in the absence of HopII, cytHsp70-1 and cpHsp70-1 may act in basal defense by facilitating folding and complex assembly of chloroplast-targeted defense factors, such as components of SA biosynthesis and transport [321]. Upon pathogen infection, HopII may switch these Hsp70s to function in degradation or disassembly of defense-promoting complexes [321].  
HopN1 is a Cys protease with experimentally confirmed proteolytic activity [328]. This effector was found to suppress PCD, ROS production, and callose deposition in plants [307,328]. The _Pst_ DC3000 _hopN1_ mutant strain with altered Cys protease catalytic triad lost its ability to suppress the hypersensitive response, indicating that the Cys protease domain is essential for hopN1 to suppress plant cell death [328]. In vitro pull-down assays identified PsbQ (protein Q in photosystem II oxygen evolution complex) as the target of HopN1 [307]. HopN1 demonstrated proteolytic activity towards PsbQ in _N. benthamiana_ thylakoids [307]. In addition, chloroplasts isolated from HopN1-expressing tomato leaves displayed reduced PSII activity [307]. In line with these observations, PsbQ-silenced tobacco plants showed reductions in bacterium-induced ROS production and cell death [307]. Therefore, HopN1 may reduce photosynthetic water splitting, oxygen production, electron transport, and ROS generation in the chloroplast by degrading PsbQ [307].