Transport intermediates (TIs) have a central role in intracellular traffic, and much effort has been directed towards defining their molecular organization. Unfortunately, major uncertainties remain regarding their true structure in living cells. To address this question, we have developed an approach based on the combination of the green fluorescent protein technology and correlative light-electron microscopy, by which it is possible to monitor an individual carrier in vivo and then take a picture of its ultrastructure at any moment of its lifecycle. We have applied this technique to define the structure of TIs operating from the Golgi apparatus to the plasma membrane, whose in vivo dynamics have been characterized recently by light microscopy. We find that these carriers are large (ranging from 0.3–1.7 μm in maximum diameter, nearly half the size of a Golgi cisterna), comprise almost exclusively tubular-saccular structures, and fuse directly with the plasma membrane, sometimes minutes after docking to the fusion site.

Genome-wide association studies of complex physiological traits and diseases consistently found that associated genetic factors, such as allelic polymorphisms or DNA mutations, only explained a minority of the expected heritable fraction. This discrepancy is known as “missing heritability”, and its underlying factors and molecular mechanisms are not established. Epigenetic programs may account for a significant fraction of the “missing heritability.” Epigenetic modifications, such as DNA methylation and chromatin assembly states, reflect the high plasticity of the genome and contribute to stably alter gene expression without modifying genomic DNA sequences. Consistent components of complex traits, such as those linked to human stature/height, fertility, and food metabolism or to hereditary defects, have been shown to respond to environmental or nutritional condition and to be epigenetically inherited. The knowledge acquired from epigenetic genome reprogramming during development, stem cell differentiation/de-differentiation, and model organisms is today shedding light on the mechanisms of (a) mitotic inheritance of epigenetic traits from cell to cell, (b) meiotic epigenetic inheritance from generation to generation, and (c) true transgenerational inheritance. Such mechanisms have been shown to include incomplete erasure of DNA methylation, parental effects, transmission of distinct RNA types (mRNA, non-coding RNA, miRNA, siRNA, piRNA), and persistence of subsets of histone marks.

Trop-2 is a calcium signal transducer that drives tumor growth. Anti-Trop-2 antibodies with selective reactivity versus Trop-2 maturation stages allowed to identify two different pools of Trop-2, one localized in the cell membrane and one in the cytoplasm. Of note, membrane-localized/functional Trop-2 was found to be differentially associated with determinants of tumor aggressiveness and distinct breast cancer subgroups. These findings candidated Trop-2 states to having an impact on cancer progression. We tested this model in breast cancer. A large, consecutive human breast cancer case series (702 cases; 8 years median follow-up) was analyzed by immunohistochemistry with anti-Trop-2 antibodies with selective reactivity for cytoplasmic-retained versus functional, membrane-associated Trop-2. We show that membrane localization of Trop-2 is an unfavorable prognostic factor for overall survival (1+ versus 0 for all deaths: hazard ratio, 1.63; P = 0.04), whereas intracellular Trop-2 has a favorable impact on prognosis, with an adjusted hazard ratio for all deaths of 0.48 (high versus low; P = 0.003). A corresponding impact of intracellular Trop-2 was found on disease relapse (high versus low: hazard ratio, 0.51; P = 0.004). Altogether, we demonstrate that the Trop-2 activation states are critical determinants of tumor progression and are powerful indicators of breast cancer patients survival.