Apocarotenoids are natural compounds derived from the oxidative cleavage of carotenoids. Particularly, C13-apocarotenoids are volatile compounds that contribute to the aromas of different flowers and fruits and are highly valued by the Flavor and Fragrance industry. So far, the chemical synthesis of these terpenoids has dominated the industry. Nonetheless, the increasing consumer demand for more natural and sustainable processes raises an interesting opportunity for bio-production alternatives. In this regard, enzymatic biocatalysis and metabolically engineered microorganisms emerge as attractive biotechnological options. The present review summarizes promising bioengineering approaches with regard to chemical production methods for the synthesis of two families of C13-apocarotenoids: ionones/dihydroionones and damascones/damascenone. We discuss each method and its applicability, with a thorough comparative analysis for ionones, focusing on the production process, regulatory aspects, and sustainability.

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SUMMARYCell growth and proliferation are pivotal for final organ and body size definition. p53-related protein kinase (Bud32/PRPK) has been identified as a protein involved in proliferation through its effects on transcription in yeast and p53 stabilization in human cell culture. However, the physiological function of Bud32/PRPK in metazoans is not well understood. In this work, we have analyzed the role of PRPK in Drosophila development. Drosophila PRPK is expressed in every tissue analyzed and is required to support proliferation and cell growth. The Prpk knockdown animals show phenotypes similar to those found in mutants for positive regulators of the PI3K/TOR pathway. This pathway has been shown to be fundamental for animal growth, transducing the hormonal and nutritional status into the protein translation machinery. Functional interactions have established that Prpk operates as a transducer of the PI3K/TOR pathway, being essential for TOR kinase activation and for the regulation of its targets (S6K and 4E-BP, autophagy and bulk endocytosis). This suggests that Prpk is crucial for stimulating the basal protein biosynthetic machinery in response to insulin signaling and to changes in nutrient availability.

Robust fermentation performance of microbial cell factories is critical for successful scaling of a biotechnological process. From shake flask cultivations to industrial-scale bioreactors, consistent strain behavior is fundamental to achieve the production targets. To assert the importance of this feature, we evaluated the impact of the yeast strain design and construction method on process scalability -from shake flasks to bench-scale fed-batch fermentations- using two recombinant Saccharomyces cerevisiae strains capable of producing β-carotene; SM14 and βcar1.2 strains. SM14 strain, obtained previously from adaptive evolution experiments, was capable to accumulate up to 21 mg/g DCW of β-carotene in 72 h shake flask cultures; while the βcar1.2, constructed by overexpression of carotenogenic genes, only accumulated 5.8 mg/g DCW of carotene. Surprisingly, fed-batch cultivation of these strains in 1L bioreactors resulted in opposite performances. βcar1.2 strain reached much higher biomass and β-carotene productivities (1.57 g/L/h and 10.9 mg/L/h, respectively) than SM14 strain (0.48 g/L/h and 3.1 mg/L/h, respectively). Final β-carotene titers were 210 and 750 mg/L after 80 h cultivation for SM14 and βcar1.2 strains, respectively. Our results indicate that these substantial differences in fermentation parameters are mainly a consequence of the exacerbated Crabtree effect of the SM14 strain. We also found that the strategy used to integrate the carotenogenic genes into the chromosomes affected the genetic stability of strains, although the impact was significantly minor. Overall, our results indicate that shake flasks fermentation parameters are poor predictors of the fermentation performance under industrial-like conditions, and that appropriate construction designs and performance tests must be conducted to properly assess the scalability of the strain and the bioprocess.

Cells extend membrane protrusions like lamellipodia and filopodia from the leading edge to sense, to move and to form new contacts. The Arp2/3 complex sustains lamellipodia formation, and in conjunction with the actomyosin contractile system, provides mechanical strength to the cell. Drosophila p53-related protein kinase (Prpk), a Tsc5p ortholog, has been described as essential for cell growth and proliferation. In addition, Prpk interacts with proteins associated to actin filament dynamics such as α-spectrin and the Arp2/3 complex subunit Arpc4. Here, we investigated the role of Prpk in cell shape changes, specifically regarding actin filament dynamics and membrane protrusion formation. We found that reductions in Prpk alter cell shape and the structure of lamellipodia, mimicking the phenotypes evoked by Arp2/3 complex deficiencies. Prpk co-localize and co-immunoprecipitates with the Arp2/3 complex subunit Arpc1 and with the small GTPase Rab35. Importantly, expression of Rab35, known by its ability to recruit upstream regulators of the Arp2/3 complex, could rescue the Prpk knockdown phenotypes. Finally, we evaluated the requirement of Prpk in different developmental contexts, where it was shown to be essential for correct Arp2/3 complex distribution and actin dynamics required for hemocytes migration, recruitment, and phagocytosis during immune response.