The role of PIWI and the miRNA machinery in Drosophila germline determination.
BACKGROUND
The germ plasm has long been demonstrated to be necessary and sufficient for germline determination, with translational regulation playing a key role in the process. Beyond this, little is known about molecular activities underlying germline determination.


RESULTS
We report the function of Drosophila PIWI, DICER-1, and dFMRP (Fragile X Mental Retardation Protein) in germline determination. PIWI is a maternal component of the polar granule, a germ-plasm-specific organelle essential for germline specification. Depleting maternal PIWI does not affect OSK or VASA expression or abdominal patterning but leads to failure in pole-plasm maintenance and primordial-germ-cell (PGC) formation, whereas doubling and tripling the maternal piwi dose increases OSK and VASA levels correspondingly and doubles and triples the number of PGCs, respectively. Moreover, PIWI forms a complex with dFMRP and DICER-1, but not with DICER-2, in polar-granule-enriched fractions. Depleting DICER-1, but not DICER-2, also leads to a severe pole-plasm defect and a reduced PGC number. These effects are also seen, albeit to a lesser extent, for dFMRP, another component of the miRISC complex.


CONCLUSIONS
Because DICER-1 is required for the miRNA pathway and DICER-2 is required for the siRNA pathway yet neither is required for the rasiRNA pathway, our data implicate a crucial role of the PIWI-mediated miRNA pathway in regulating the levels of OSK, VASA, and possibly other genes involved in germline determination in Drosophila.
Megosh
Heather B
HB
Department of Cell Biology and Duke University Medical Center, Durham, North Carolina 27705, USA.
Cox
Daniel N
DN
Campbell
Chris
C
Lin
Haifan
H
D000818:Animals; D060565:Argonaute Proteins; D019521:Body Patterning / Q000502:physiology; D002454:Cell Differentiation; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism; D004330:Drosophila / Q000196:embryology* / Q000235:genetics / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D004625:Embryo, Nonmammalian / Q000166:cytology / Q000378:metabolism; D047108:Embryonic Development / Q000502:physiology; D051860:Fragile X Messenger Ribonucleoprotein 1 / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D018507:Gene Expression Regulation, Developmental; D005854:Germ Cells / Q000166:cytology / Q000378:metabolism*; D035683:MicroRNAs / Q000502:physiology*; D011506:Proteins / Q000502:physiology*; D020365:RNA Helicases / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D034741:RNA, Small Interfering / Q000378:metabolism; D034743:RNA-Induced Silencing Complex; D043244:Ribonuclease III
D060565:Argonaute Proteins; D053487:DEAD-box RNA Helicases; D029721:Drosophila Proteins; D051860:Fragile X Messenger Ribonucleoprotein 1; D035683:MicroRNAs; D011506:Proteins; D020365:RNA Helicases; D034741:RNA, Small Interfering; D034743:RNA-Induced Silencing Complex; D043244:Ribonuclease III; C091675:osk protein, Drosophila; C116498:piwi protein, Drosophila; C478069:DCR-2 protein, Drosophila; C515033:Dcr-1 protein, Drosophila; C088269:vas protein, Drosophila