Little is known of the molecular mechanisms whereby spermatogonia, mitotic germ cells of the testis, self-renew and differentiate into sperm 1,2 . Here we show that Zfp145, encoding the transcriptional repressor Plzf, has a crucial role in spermatogenesis. Zfp145 expression was restricted to gonocytes and undifferentiated spermatogonia and was absent in tubules of W/W v mutants that lack these cells. Mice lacking Zfp145 underwent a progressive loss of spermatogonia with age, associated with increases in apoptosis and subsequent loss of tubule structure but without overt differentiation defects or loss of the supporting Sertoli cells. Spermatogonial transplantation experiments revealed a depletion of spermatogonial stem cells in the adult. Microarray analysis of isolated spermatogonia from Zfp145-null mice before testis degeneration showed alterations in the expression profile of genes associated with spermatogenesis. These results identify Plzf as a spermatogoniaspecific transcription factor in the testis that is required to regulate self-renewal and maintenance of the stem cell pool.

The molecular mechanisms that regulate coordinated and colinear activation of Hox gene expression in space and time remain poorly understood. Here we demonstrate that Plzf regulates the spatial expression of the AbdB HoxD gene complex by binding to regulatory elements required for restricted Hox gene expression and can recruit histone deacetylases to these sites. We show by scanning forced microscopy that Plzf, via homodimerization, can form DNA loops and bridge distant Plzf binding sites located within HoxD gene regulatory elements. Furthermore, we demonstrate that Plzf physically interacts with Polycomb proteins on DNA. We propose a model by which the balance between activating morphogenic signals and transcriptional repressors such as Plzf establishes proper Hox gene expression boundaries in the limb bud.

Here we report the synthesis of PLGA/DOXO-core Au-branched shell nanostructures (BGNSHs) functionalized with a human serum albumin/indocyanine green/folic acid complex (HSA-ICG-FA) to configure a multifunctional nanotheranostic platform. First, branched gold nanoshells (BGNSHs) were obtained through a seeded-growth surfactant-less method. These BGNSHs were loaded during the synthetic process with the chemotherapeutic drug doxorubicin, a DNA intercalating agent and topoisomerase II inhibitior. In parallel, the fluorescent near-infrared (NIR) dye indocyanine green (ICG) was conjugated to the protein human serum albumin (HSA) by electrostatic and hydrophobic interactions. Subsequently, folic acid was covalently attached to the HSA-ICG complex. In this way, we created a protein complex with targeting specificity and fluorescent imaging capability. The resulting HSA-ICG-FA complex was adsorbed to the gold nanostructures surface (BGNSH-HSA-ICG-FA) in a straightforward incubation process thanks to the high affinity of HSA to gold surface. In this manner, BGNSH-HSA-ICG-FA platforms were featured with multifunctional abilities: the possibility of fluorescence imaging for diagnosis and therapy monitoring by exploiting the inherent fluorescence of the dye, and a multimodal therapy approach consisting of the simultaneous combination of chemotherapy, provided by the loaded drug, and the potential cytotoxic effect of photodynamic and photothermal therapies provided by the dye and the gold nanolayer of the hybrid structure, respectively, upon NIR light irradiation of suitable wavelength. The combination of this trimodal approach was observed to exert a synergistic effect on the cytotoxicity of tumoral cells in vitro. Furthermore, FA was proved to enhance the internalization of nanoplatform. The ability of the nanoplatforms as fluorescence imaging contrast agents was tested by preliminary analyzing their biodistribution in vivo in a tumor-bearing mice model.