PIAS (protein inhibitor of activated STAT) proteins interact with and modulate the activities of various transcription factors. In this work, we demonstrate that PIAS proteins x␣, x␤, 1, and 3 interact with the small ubiquitin-related modifier SUMO-1 and its E2 conjugase, Ubc9, and that PIAS proteins themselves are covalently modified by SUMO-1 (sumoylated). PIAS proteins also tether other sumoylated proteins in a noncovalent fashion. Furthermore, recombinant PIASx␣ enhances Ubc9-mediated sumoylation of the androgen receptor and c-Jun in vitro. Importantly, PIAS proteins differ in their abilities to promote sumoylation in intact cells. The ability to stimulate protein sumoylation and the interaction with sumoylated proteins are dependent on the conserved PIAS RING finger-like domain. These functions are linked to the activity of PIASx␣ on androgen receptor-dependent transcription. Collectively, our results imply that PIAS proteins function as SUMO-1-tethering proteins and zinc finger-dependent E3 SUMO protein ligases, and these properties are likely to explain their ability to modulate the activities of various transcription factors.Members of the recently identified PIAS (protein inhibitor of activated STAT) protein family have been found to interact with several distinct nuclear proteins. PIAS1 and PIAS3 bind to STAT1 and STAT3, respectively, and inhibit their action (4, 28). PIASx␣/ARIP3 (androgen receptor [AR]-interacting protein 3) was first characterized as an AR-interacting protein, and it modulates the transcriptional activity of the receptor (34). Other PIAS proteins have recently been demonstrated to function as coregulators for AR and other steroid receptors (25, 57). PIASx␤/Miz1 (Msx-interacting zinc finger) associates with a homeodomain-containing Msx2 protein (61), and GBP (Gu/RNA helicase II-binding protein), which is nearly identical to PIAS1, interacts with Gu/RNA II-helicase (59). More recently, PIAS proteins have been identified in many yeast two-hybrid screens, including PIASx␣/ARIP3 from its interaction with mouse disabled 2 (mDab2) (3) and DJ-1 protein (54), PIAS1 from its interaction with p53 (9), and PIAS3 from its interaction with high-mobility-group protein HMGI-C (63) and zinc finger protein Gfi-1 (43). In addition to PIASx (PIASx␣/ARIP3 and PIASx␤/Miz1), SUMO-1 (small ubiquitinrelated modifier 1) was identified as a p73␣-interacting protein by Minty et al. (32), who suggested that PIASx was isolated via the interaction with Smt3p (yeast SUMO) covalently linked to p73␣. We have also detected SUMO-1 as a major PIASx␣/ ARIP3-interacting protein in yeast (unpublished results).Members of the SUMO protein family, also known as Sentrin, GMP1, PIC1, and Ubl1 (31, 37, 62), are present in protozoa, metazoa, plants, and fungi. SUMO proteins from metazoa can be divided into two families: the SUMO-1 family and the SUMO-2 and -3 family (31, 37, 62). SUMO-2 and SUMO-3 are very similar at the amino acid level (97% identity for the human proteins), but they are only ϳ50% identical to SUMO-1. SUMO-1 is...

The chromatoid body is a perinuclear, cytoplasmic cloud-like structure in male germ cells whose function has remained elusive. Here we show that the chromatoid body is related to the RNA-processing body of somatic cells. Dicer and components of microRNP complexes (including Ago proteins and microRNAs) are highly concentrated in chromatoid bodies. Furthermore, we show that Dicer interacts with a germ cell-specific chromatoid body component, the RNA helicase MVH (mouse VASA homolog). Thus, chromatoid bodies seem to operate as intracellular nerve centers of the microRNA pathway. Our findings underscore the importance of posttranscriptional gene regulation and of the microRNA pathway in the control of postmeiotic male germ cell differentiation.Argonaute ͉ microRNA ͉ spermatogenesis ͉ RNA processing

The chromatoid body, a unique cloud-like structure of male germ cells, moves dynamically in the cytoplasm of haploid spermatids, but its function has remained elusive for decades. Recent findings indicate that microRNA and RNA-decay pathways converge to the chromatoid body. This highly specialized structure might function as an intracellular focal domain that organizes and controls RNA processing in male germ cells.

MIWI catalytic activity is required for spermatogenesis, indicating that piRNA-guided cleavage is critical for germ cell development. To identify meiotic piRNA targets, we augmented the mouse piRNA repertoire by introducing a human meiotic piRNA cluster. This triggered a spermatogenesis defect by inappropriately targeting the piRNA machinery to mouse mRNAs essential for germ cell development. Analysis of such de novo targets revealed a signature for pachytene piRNA target recognition. This enabled identification of both transposable elements and meiotically expressed protein-coding genes as targets of native piRNAs. Cleavage of genic targets began at the pachytene stage and resulted in progressive repression through meiosis, driven at least in part via the ping-pong cycle. Our data support the idea that meiotic piRNA populations must be strongly selected to enable successful spermatogenesis, both driving the response away from essential genes and directing the pathway toward mRNA targets that are regulated by small RNAs in meiotic cells.

No abstract