Updated project metadata. The chloroplast signal recognition particle 54 kDa (CpSRP54) is a member of the CpSRP pathway targeting proteins to thylakoid membranes. In plants, CpSRP54 has a dual function and is involved in both co-translational and post-translational insertion of thylakoid membrane proteins like subunits of photosynthetic complexes and light harvesting antenna proteins, respectively. In green microalgae, CpSRP54 seems to function only in the post-translational part of the CpSRP pathway. The effects of loss of CpSRP54 in the diatom model species Phaeodactylum tricornutum support a role in co-translational targeting of proteins to thylakoid membranes, but not in the post-translational CpSRP pathway. Lack of CpSRP54 does not have a negative effect on the content of light harvesting antenna proteins and pigments, but causes lower levels of a selection of PSII, PSI, Cytb6f and ATP synthase subunits. Cpsrp54 lines also display decreased photophysiological fitness, stronger induction of photoprotective mechanisms and lower growth rates compared to wild type when exposed to increased light intensities. Mechanisms alleviating the loss of CpSRP54, involving upregulation of chaperones, might explain the still relatively mild phenotype of the cpsrp54 mutants.