Direkt zum Inhalt
Merck
  • D-Aspartate Upregulates DAAM1 Protein Levels in the Rat Testis and Induces Its Localization in Spermatogonia Nucleus.

D-Aspartate Upregulates DAAM1 Protein Levels in the Rat Testis and Induces Its Localization in Spermatogonia Nucleus.

Biomolecules (2020-05-02)
Massimo Venditti, Alessandra Santillo, Sara Falvo, Maria Maddalena Di Fiore, Gabriella Chieffi Baccari, Sergio Minucci
ZUSAMMENFASSUNG

Cell differentiation during spermatogenesis requires a proper actin dynamic, regulated by several proteins, including formins. Disheveled-Associated-Activator of Morphogenesis1 (DAAM1) belongs to the formins and promotes actin polymerization. Our results showed that oral D-Aspartate (D-Asp) administration, an excitatory amino acid, increased DAAM1 protein levels in germ cells cytoplasm of rat testis. Interestingly, after the treatment, DAAM1 also localized in rat spermatogonia (SPG) and mouse GC-1 cells nuclei. We provided bioinformatic evidence that DAAM1 sequence has two predicted NLS, supporting its nuclear localization. The data also suggested a role of D-Asp in promoting DAAM1 shuttling to the nuclear compartment of those proliferative cells. In addition, the proliferative action induced by D-Asp is confirmed by the increased levels of PCNA, a protein expressed in the nucleus of cells in the S phase and p-H3, a histone crucial for chromatin condensation during mitosis and meiosis. In conclusion, we demonstrated, for the first time, an increased DAAM1 protein levels following D-Asp treatment in rat testis and also its localization in the nucleus of rat SPG and in mouse GC-1 cells. Our results suggest an assumed role for this formin as a regulator of actin dynamics in both cytoplasm and nuclei of the germ cells.

MATERIALIEN
Produktnummer
Marke
Produktbeschreibung

Sigma-Aldrich
Dulbeccos Phosphatgepufferte Kochsalzlösung, Modified, without calcium chloride and magnesium chloride, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
Albumin aus Rinderserum, lyophilized powder, ≥96% (agarose gel electrophoresis)
Sigma-Aldrich
Trypsin-EDTA-Lösung, 10 ×, sterile-filtered, BioReagent, suitable for cell culture, 5.0 g porcine trypsin and 2 g EDTA, 4Na per liter of 0.9% sodium chloride
Sigma-Aldrich
Anti-Phospho-Histon H3 (Ser10)-Antikörper, Mitosemarker, Upstate®, from rabbit
Sigma-Aldrich
IgG aus Kaninchenserum, reagent grade, ≥95% (SDS-PAGE), essentially salt-free, lyophilized powder
Sigma-Aldrich
Anti-Kaninchen-IgG-Antikörper der Ziege, HRP-konjugiert, 1 mg/mL, Upstate®
Sigma-Aldrich
Paraformaldehyd, powder, 95%
Sigma-Aldrich
D-Asparaginsäure, ReagentPlus®, 99%
Sigma-Aldrich
Anti-Histon H3-Antikörper, 0.5 mg/mL, Upstate®
Sigma-Aldrich
Anti-Mouse IgG (H+L), CF 568 antibody produced in goat, ~2 mg/mL, affinity isolated antibody
Sigma-Aldrich
Anti-DAAM1 antibody produced in rabbit, affinity isolated antibody