Chloroplasts are specialized plastids that host photosynthetic reactions in green tissues. These highly specialized organelles have evolved from an endosymbiotic event, where a photosynthetic prokaryotic organism was engulfed by a proeukaryotic cell. During evolution and in parallel with the increasing complexity of multicellular photosynthetic organisms, these semi-autonomous organelles have diversified into a wide group of plastids to adapt to specific cellular and tissular functions. In plants chloroplasts are inherited as non-photosynthetic plastids, the proplastids, which can differentiate into chloroplasts in green tissues or other specialized types of plastids, such as chromoplasts or amyloplast, following precise developmental programmes and signals (Fig 1). Moreover, plastids can interconvert into other types in response to external and internal signals and cues.

Fig. 1. Plastids diversity, differentiation and interconversion.

After the endosymbiotic event and during evolution, the plastids lost or transferred most of its genes to the host genome, becoming semi-autonomous since they depend on the nucleus for their development and function. The division of genetic information between the nuclear and the plastid genomes requires a precise coordination that is achieved through a complex signalling, from the nucleus to the plastids (anterograde) and from plastids to the nucleus (retrograde). Although chloroplast biogenesis is under nuclear control, retrograde signals are required for the successful completion of the process.

Our research seeks to identify new nuclear factors involved in chloroplast biogenesis and molecular mechanisms that determine chloroplast development in specific cell lineages.

Research lines

Transcriptional regulation of chloroplast biogénesis

In dark-grown -etiolated- seedlings, chloroplast biogenesis is tightly suppressed by transcriptional and post-transcriptional mechanisms to prevent over-accumulation of chlorophyll intermediates that can cause damage upon light exposure (Fig 2).  Exposure to light initiates chloroplast biogenesis that can be divided into two distinct phases, assembly and photosynthesis (Fig 2). The assembly phase starts with light exposure, which is sensed by photoreceptors, such as phytochromes and cryptochromes. Active phytochromes are translocated to the nucleus degrading PIFs and EIN3, resulting in the stabilisation of transcription factors such as HY5 and the GLKs activating transcription of a large number of genes encoding chloroplast proteins. These proteins are imported into the developing etioplasts, and photosynthetic complexes begin to assemble, starting the transition to photosynthetically active chloroplasts. The second phase, photosynthesis, depends on plastid retrograde signalling (RS) mediated mainly by GUN1 which further boosts nuclear transcription of Photosynthetic Associated Nuclear Genes (PhANGs).

Fig. 2. Overview of chloroplast development. Chloroplast biogenesis starts from proplastids, or the intermediate etioplast in cotyledons of dark-grown seedlings. Nuclear transcription of PhANGs is repressed by PIFs in cooperation with other proteins like EIN3. GUN1-dependent retrograde signal is present in proplastids and etioplasts, repressing chloroplast biogenesis. Perception of light by photoreceptors triggers chloroplast biogenesis and the transition to photoautotrophic growth. Transcription factors, like HY5, are stabilised and as a result, there is an increase in the expression of PhANGs, and plastids proteins are imported to plastids to start the assembly of plastid protein complexes. The retrograde signal is still present in the developing chloroplasts, holding nuclear transcription and full development. The increase in plastid transcription and the correct progression of plastid development, during the assembly phase, degrades GUN1 and boosts GLKs and PhANGs transcription, leading to fully functional chloroplast and photoautotrophic growth. (Adapted from Hernández-Verdeja, Journal of Experimental Botany, 2025).

Our results revealed that the plastid retrograde signal is present in proplastids and etioplasts and starts to fade during the assembly phase repressing expression of PhANGs. With correct chloroplast development, GUN1 is degraded lifting the repression of nuclear transcription factors and PhANGs expression is boosted. We hypothesise that the plastid retrograde signal regulates more transcriptional regulators during chloroplast development and this project aims to identify and characterise these transcription factors.

In most plants lineages, some cells and tissues, such as the root, or leaf bundle sheath cells (BSC), do not develop photosynthetic chloroplast even when exposed to light. The signalling pathways regulating the transition to chloroplasts in specific cells and tissues remain mostly unknown yet this is particularly relevant for evolutionary transitions to more efficient photosynthetic types. Some plant lineages, such as C₄ and C₂, have evolved carbon concentrating mechanisms (CCM) to minimise photorespiration costs and boost net carbon assimilation.

C₂ photosynthesis, also termed the glycine shuttle or photorespiratory CO₂ pump. C₂ plants spatially separate photorespiration by shuttling the photorespiratory product glycine from mesophyll to bundle sheath cells where it releases, concentrates, and re-assimilates CO₂ to bolster photosynthetic efficiency. In contrast to C₃ plants, bundle sheath cells of C₂ plants have abundant chloroplasts to re-assimilates the CO₂ from the decarboxylation of glycine (Fig 3).

Fig. 3. C2 photosynthesis. (a) Schematic of C3 and C2 photosynthesis. In C2 plants the glycine produced by the Rubisco oxygenation diffuses to the BSC mitochondria to be metabolized to serine and CO2 in the mitochondria by glycine decarboxylase. CO2 released is refixed by BSC chloroplasts. (b) Representative images of bundle sheath organelle ultrastructure of C2 D. tenuifolia and C3 Diplotaxis viminea. BSC, bundle sheath cell; ch, chloroplast; asteriks, mitochondria. Scale bars = 800 nm. (c) BSC chloroplasts (left), and mitochondria (right) number per um of BSC-VS shared cell wall.