Human Pumilio-2 is expressed in embryonic stem cells and germ cells and interacts with DAZ (Deleted in AZoospermia) and DAZ-Like proteins

Abstract

Early in development, a part of the embryo is set aside to become the germ cell lineage that will ultimately differentiate to form sperm and eggs and transmit genetic information to the next generation. Men with deletions encompassing the Y-chromosome DAZ genes have few or no germ cells but are otherwise healthy, indicating they harbor specific defects in formation or maintenance of germ cells. A DAZ homolog, DAZL (DAZ-Like), is found in diverse organisms, including humans and is required for germ cell development in males and͞or females. We identified proteins that interact with DAZ proteins to better understand their function in human germ cells. Here, we show that PUM2, a human homolog of Pumilio, a protein required to maintain germ line stem cells in Drosophila and Caenorhabditis elegans, forms a stable complex with DAZ through the same functional domain required for RNA binding, protein-protein interactions and rescue of Pumilio mutations in flies. We also show that PUM2 is expressed predominantly in human embryonic stem cells and germ cells and colocalizes with DAZ and DAZL in germ cells. These data implicate PUM2 as a component of conserved cellular machinery that may be required for germ cell development.A ll stem cells have potential to differentiate or proliferate mitotically. These potentials must be balanced for the stem cell population to be maintained. If differentiation exceeds proliferation, the stem cell population is not maintained. Evidence in humans suggests that the DAZ genes function early in the germ line stem cells. Men with deletions encompassing the Y-chromosome DAZ gene cluster have defects in spermatogenesis that are detected initially in the stem cell population. These men frequently lack all germ cells, including the spermatogonial stem cells, and only somatic cells are present in testicular tissue (1-3). In addition, expression of the DAZ gene and its ancestral, autosomal homolog, DAZL, only occurs in germ cells; DAZ is expressed in males, and DAZL is expressed in males and females (4, 5). Evidence from model organisms also suggests the DAZ genes function in germ cell maintenance. The Xenopus homolog of DAZ, Xdazl, is expressed in a region of the early oocyte that contains the germ plasm that is required for formation and maintenance of the germ cell lineage (6, 7). Inhibition of Xdazl leads to loss of the primordial germ cells (6, 7). Finally, the DAZ homolog in mice, Dazl, is most abundantly expressed in premeiotic germ cells; disruption of this gene causes depletion of germ cells beginning prenatally (8-10). DAZ and DAZL homologs may function interchangeably as suggested by the observation that a human DAZ transgene can partially rescue a mouse Dazl mutation (11). To shed light on how DAZ genes might function in human germ cells, we sought to identify proteins that interact with DAZ proteins. Materials and MethodsTwo-Hybrid Screening of DAZ-Interacting Proteins. The yeast two-hybrid system was used to identify proteins that interact with a DAZ:GAL4 DNA-binding domain fusion prot...

Regulated Pumilio-2 binding controls RINGO/Spy mRNA translation and CPEB activation

“…The significance of these interactions, however, has not been established. Pum2, on the other hand, interacts with human and mouse DAZ and DAZL (Moore et al 2003). Similarly, in Drosophila, Pum2 interacts with Boule, the DAZ homolog in this species .…”

“…The interaction of DAZL with RINGO/Spy RNA is perhaps not surprising given that DAZL and Pum2 can bind one another (Moore et al 2003). DAZL also binds ePAB, a Xenopus embryonic PABP that stabilizes poly(A) tails on RNAs during maturation and early embryogenesis (Voeltz et al 2001;Collier et al 2005).…”

Metazoan oocyte and early embryo development program: a progression through translation regulatory cascades

“…One such negative regulator, the Pumilio/PBE complex, functions to repress translation, presumably through its interactions with the DAZL/ePABP complex (Fig. 1, green box; Moore et al 2003;Padmanabhan and Richter 2006). Conversely, in the absence of negative elements such as the PBE, as in Figure 3 of the article by Padmanabhan and Richter (2006), translation was activated without a need for meiotic activation or subsequent maturation signals.…”

“…Second, xPumilio can interact with CPEB through its PUF RNA-binding motif and may thereby impose an additional constraint on CPEB-mediated translation regulation (Nakahata et al 2003). This limitation may involve a mechanism similar to the one discovered by Padmanabhan and Richter (2006), where xPumilio bound to the xPB site interacts in cis with additional 3Ј-UTR element regulatory complexes such as CPEB or DAZL/ ePABP (Moore et al 2003;Nakahata et al 2003). Repression of cyclin B1 mRNA is released upon maturation through activation of CPEB and loss of xPumilio binding to the mRNA and to CPEB; as a consequence, translation proceeds through a CPE-mediated polyadenylation mechanism where loss of repression requires ePABP recruitment Nakahata et al 2003).…”

Pumilio Puf domain RNA-binding proteins in Arabidopsis

“…The data on and thus appeared to be required for maintenance of the human germ cell lineage. 22 In yeast, Puf3p is involved in mitochondrial biogenesis as it preferentially binds to the mRNAs of nuclear-encoded mitochondrial proteins. 23 It also contributes to mitochondrial motility during budding from the mother cell by linking the mitochore, a protein complex implicated in linking mitochondria to actin filaments, with the Arp2/3 complex, which generates the force needed for actin dependent mitochondria movement.…”