For the last 20 years, the Kremer lab has had a broad interest in adenovirus biology. Adenoviruses are nonenveloped, double-stranded DNA pathogens with a genome size of 28 – 42 kb. To date, ~70 human and > 150 nonhuman adenovirus types have been described. We focus on human and canine adenoviruses (CAV-2) vectors.
OUR PRINCIPLE AREAS OF INTEREST ARE:
1.Understanding the function of CAR, the coxsackievirus adenovirus receptor, in the healthy and disease brain, and during adenovirus trafficking.
As the name suggests, CAR was originally identified as an attachment molecule for coxsackie B viruses and some adenoviruses, including canine adenovirus type 2 (CAV-2) vectors, whose transduction efficacy is CAR dependent. CAR is a single-pass transmembrane protein belonging to the CTX subfamily of the Ig-super family. CAR function is best characterized in epithelial cells where it participates to the maintenance of tight junctions. Although a role for CAR in brain development has been hypothesized and a function in regulating axonal growth reported, its role in the adult brain is largely uncharacterized. In this context, understanding CAR’s biological function, as well as the impact of its loss of function and distribution in the CNS is indispensable for both fundamental and applied neurobiology studies.
2.Understanding how mononuclear phagocytes, in particular dendritic cells, process and respond to immune-complexed adenoviruses (IC-Ad)
The nearly ubiquitous presence of anti-adenovirus antibodies and memory T cells is due to recurrent and cross-reacting adenovirus infections during childhood. Thus, when Ad-based vectors are used in a host with memory immunity this creates an environment close to a secondary infection in the clinic.
How do IC-Ads and other host factors impact the maturation of antigen-presenting cells (APC)? Our underlying questions encompass the entry and signaling pathways and how the IC-Ads traffic in APC. In our studies we use human dendritic cells to study the interaction of IC-HAd5 and innate immune sensors. Dendritic cells are the major APC in blood and tissue and play a pivotal role in the sensing of infection and activation and re-activation of the adaptive immune response.
IC-Ads triggers several innate immune sensors and induce DC maturation (the virus alone is far less potent). Our aim is to understand the interactions of the IC-Ads during the initial activation and death of DC. Our studies contribute to both fundamental and applied questions in adenovirus biology, vaccination and gene therapy.
3.Optimizing CAV-2 vector for gene transfer to the CNS
The Kremer labs created replication-defective and helper-dependent CAV-2 vectors. CAV-2 vectors allow preferential gene transfer to neurons and widespread distribution after injection into the brain. We and others have used them to explore, treat and understand the healthy and diseased CNS.
Composition of the sub-group: Florence Rage, Johann Soret, Pauline Duc et Audrey Moisan.
Understanding the molecular mechanisms of Spinal Muscular Atrophy (SMA)
Spinal Muscular Atrophy (SMA), a hereditary disease, results from degeneration of motor neurons leading to severe muscular atrophy. This disease is caused by the deletion and/or mutation of the SMN1 gene. Due to its crucial role in the biogenesis of spliceosome components, SMN deficiency is correlated to numerous splicing alterations. At the cytoplasmic level, given the high polarity of motor neurons, multiple other molecular mechanisms are impaired in this disease, including mRNA transport along axons of motor neurons and the formation and the maintenance of neuromuscular junctions (NMJs).
In our projects, we are using human induced pluripotent stem cells (hIPSCs) from healthy and SMA individuals to obtain motor neurons and study :
- The splicing defects of mRNAs in SMN deficient cells.
- The localization and local translation of mRNAs along axons of motor neurons using the MS2 and SunTag systems, integrated in target genes by the CRISPR-Cas9 technology.
- The SMN-interacting protein network in control and SMA MNs and myoblasts derived from iPSCs by combining CRISPR-Cas9 technology and biotin proximity-dependent labeling (BioID).
- The formation, development and maintenance of mature and functional human NMJs using microfluidic devices coupled with microelectrode array (MEA) in an interdisciplinary collaboration with Benoît Charlot (IES) and Gilles Carnac (Phymedexp) (Duc et al., 2021 Lab Chip, doi 10.1039/d1lc00497b).
- Finally, in collaboration with Eran Perlson (Tel Aviv University, Israel), we are also developing 24-well plate devices, combined with MEA, in order to recapitulate NMJs from SMA and ALS patients and to perform drug assays with personalized medicine approach.