Thank ORP9 for FFAT: With endosomal ORP10, it’s fission accomplished!

Heterogeneity in endosomal membrane phospholipid content is emerging as a regulator of endocytic trafficking pathways. Kawasaki et al. (2021. J. Cell. Biol.https://doi.org/10.1083/jcb.202103141) demonstrate exchange of endosomal PI4P for PS by ORP10 at ER–endosome contact sites, with the consequent recruitment of endosomal fission factors.

Endocytic Trafficking Promotes Vacuolar Enlargements for Fast Cell Expansion Rates in Plants

The vacuole has a space-filling function, allowing a particularly rapid plant cell expansion with very little increase in cytosolic content (Loefke et al., 2015; Scheuring et al., 2016; Duenser et al., 2019). Despite its importance for cell size determination in plants, very little is known about the mechanisms that define vacuolar size. Here we show that the cellular and vacuolar size expansions are coordinated. By developing a pharmacological tool, we enabled the investigation of membrane delivery to the vacuole during cellular expansion. Counterintuitively, our data reveal that endocytic trafficking from the plasma membrane to the vacuole is enhanced in the course of rapid root cell expansion. While this "compromise" mechanism may theoretically at first decelerate cell surface enlargements, it fuels vacuolar expansion and, thereby, ensures the coordinated augmentation of vacuolar occupancy in dynamically expanding plant cells.

Endocytic Protein Defects in the Neural Crest Cell Lineage and Its Pathway Are Associated with Congenital Heart Defects

Endocytic trafficking is an under-appreciated pathway in cardiac development. Several genes related to endocytic trafficking have been uncovered in a mutagenic ENU screen, in which mutations led to congenital heart defects (CHDs). In this article, we review the relationship between these genes (including LRP1 and LRP2) and cardiac neural crest cells (CNCCs) during cardiac development. Mice with an ENU-induced Lrp1 mutation exhibit a spectrum of CHDs. Conditional deletion using a floxed Lrp1 allele with different Cre drivers showed that targeting neural crest cells with Wnt1-Cre expression replicated the full cardiac phenotypes of the ENU-induced Lrp1 mutation. In addition, LRP1 function in CNCCs is required for normal OFT lengthening and survival/expansion of the cushion mesenchyme, with other cell lineages along the NCC migratory path playing an additional role. Mice with an ENU-induced and targeted Lrp2 mutation demonstrated the cardiac phenotype of common arterial trunk (CAT). Although there is no impact on CNCCs in Lrp2 mutants, the loss of LRP2 results in the depletion of sonic hedgehog (SHH)-dependent cells in the second heart field. SHH is known to be crucial for CNCC survival and proliferation, which suggests LRP2 has a non-autonomous role in CNCCs. In this article, other endocytic trafficking proteins that are associated with CHDs that may play roles in the NCC pathway during development, such as AP1B1, AP2B1, FUZ, MYH10, and HECTD1, are reviewed.

Dysregulation of the AP2M1 phosphorylation cycle by LRRK2 impairs endocytosis and leads to dopaminergic neurodegeneration

Mutations in the kinase LRRK2 and impaired endocytic trafficking are both implicated in the pathogenesis of Parkinson’s disease (PD). Expression of the PD-associated LRRK2 mutant in mouse dopaminergic neurons was shown to disrupt clathrin-mediated endocytic trafficking. Here, we explored the molecular mechanism linking LRRK2 to endocytosis and found that LRRK2 bound to and phosphorylated the μ2 subunit of the adaptor protein AP2 (AP2M1), a core component of the clathrin-mediated endocytic machinery. Analysis of human SH-SY5Y cells and mouse neurons and tissues revealed that loss of LRRK2 abundance or kinase function resulted in decreased phosphorylation of AP2M1, which is required for the initial formation of clathrin-coated vesicles (CCVs). In contrast, overexpression of LRRK2 or expression of a Parkinson’s disease–associated gain-of-function mutant LRRK2 (G2019S) inhibited the uncoating of AP2M1 from CCVs at later stages and prevented new cycles of CCV formation. Thus, the abundance and activity of LRRK2 must be calibrated to ensure proper endocytosis. Dysregulated phosphorylation of AP2M1 from the brain but not thyroid tissues of LRRK2 knockout and G2019S-knockin mice suggests a tissue-specific regulatory mechanism of endocytosis. Furthermore, we found that LRRK2-dependent phosphorylation of AP2M1 mediated dopaminergic neurodegeneration in a Drosophila model of PD. Together, our findings provide a mechanistic link between LRRK2, AP2, and endocytosis in the pathogenesis of PD.

RAB5A expression is a predictive biomarker for trastuzumab emtansine in breast cancer

PURPOSE: Targeted therapeutics strongly depends on validated biomarkers in order to select patients most likely to benefit from the treatment. HER2 serves as a predictive biomarker for HER2-targeted tyrosine kinase inhibitors and monoclonal antibodies. HER2 may, however, also be utilized as a transport gate for delivery of cytotoxic agents into the cell, such as for HER2-targeted antibody drug conjugates (ADCs; e.g. trastuzumab emtansine (T-DM1)). The predictive biomarkers for such ADCs may be more complex, also reflecting the intracellular transport. METHODS: Five HER2-positive breast and ovarian cancer cell lines were evaluated with respect to T-DM1 sensitivity and correlated to the expression levels of proteins involved in endocytic trafficking including RAB4A, RAB5A and RAB11A, with possible impact on ADC pharmacology. The results were confirmed in a clinical cohort consisting of patients from the adaptive breast cancer clinical trial I-SPY2 where pathological complete response (pCR) was correlated to the RNA expression level of RAB4A, RAB5A and RAB11A. A subset of the clinical KAMILLA trial including 19 patients was used as a verification cohort where semi-quantitative IHC of RAB5A was correlated to progression free survival (PFS). RESULTS: The early endosome marker RAB5A, was found to correlate positively to T-DM1 sensitivity in the cell line panel. Correlation between RAB5A expression and T-DM1 sensitivity (pCR) was confirmed in patients treated with trastuzumab emtansine/pertuzumab in the I-SPY2 trial, but not in the trastuzumab/paclitaxel control arm. The clinical correlation was verified in the patients from the KAMILLA trial where semi- quantitative RAB5A IHC staining correlated significantly positive to PFS. CONCLUSION: The present results indicate that RAB5A is a predictive biomarker for T-DM1 and outline, for the first time, proteins involved in endocytic trafficking as predictive biomarkers for ADCs.