Unit 11 Anterior Posterior Axis Specification in Drosophila Drosophila cleavage creates the blastoderm Initially nuclei divide in the interior many then migrate to the periphery to form the syncytial blastoderm Eventually membrane surround these nuclei to form the cellular blastoderm See Campbell 6e Fig 21 11 Cellularization involves deepening of membrane inpocketings around nuclei Baskets of microtubules surround nuclei and actin concentrates at the deepest points of inpocketing Cadherin containing junctions also track with the actin membrane furrows not shown Gilbert 10e Fig 6 4 A P Patterning Overview Gilbert 10e Fig 6 7 Segmental divisions and segmental identity are established in the embryo Islands of cells imaginal discs in the larva give rise to adult tissues that reflect these segmental identities A P Patterning Overview A P polarity depends on maternal effect genes Hunchback protein gradient Hunchback a gap gene gradient results from both maternal and zygotic hunchback Gap genes Gilbert 10e Fig 6 18 Other gap genes are zygotically expressed as are subsequent genes A P Patterning Overview Pair rule genes are expressed in 7 stripe patterns the syncytial blastoderm embryo Pair rule genes Segment polarity genes are expressed in 14stripe patterns in the cellular blastoderm embryo Gilbert 10e Fig 6 18 Homeotic genes establish segment identity A P Patterning Nobel Laureates WSJ 1995 Denticle bands allow identification of larval segments and polarity in Drosophila Denticle bands Gilbert 10e Fig 6 6 Denticle bands allow identification of segmentation mutants position and number of bands allows us to infer what is missing Courtesy FlyMove Maternal polarity centers can be identified via cytoplasm transfer anterior posterior normal fertilized egg prick anterior end allow cytoplasm to ooze out transplant posterior cytoplasm from donor into anterior end allow to develop to larval stage normal larva double posterior larva Conclusion there appear to be localized determinants or activities at both the anterior and posterior ends of the fertilized egg adapted from Alberts et al 3e Fig 21 58 Maternal polarity mutants fall into specific classes normal torso bicoid nanos APT PT A T AP Normal larvae have terminal T anterior A and posterior P structures adapted from Alberts et al 3e Fig 21 53 Effects of absence of Bicoid protein Gilbert 10e Fig 6 25 bicoid is key for anterior and nanos for posterior structures bicoid mRNA is localized at the anterior nanos at the posterior After fertilization the proteins adopt a graded distribution Gilbert 10e Fig 6 20 bicoid and nanos mRNAs localize during oogenesis Gilbert 10e Fig 6 8 Maternal mRNAs come from nurse cells nurse cells oocyte ring canals Nurse cells send mRNAs from the mother to the oocytes They are connected to the oocyte via actin rich structures called ring canals bicoid is key for anterior and nanos for posterior structures bicoid mRNA is localized at the anterior nanos at the posterior After fertilization the proteins adopt a graded distribution Gilbert 10e Fig 6 20 Bicoid protein adopts a gradient bicoid mRNA is tightly localized at the anterior bicoid protein adopts a gradient bcd mRNA bcd protein bicoid protein regulates caudal translation bicoid regulates caudal translation Gilbert 10e Fig 6 24 Zygotic hunchback mRNA is regulated by bcd bicoid protein is graded at the anterior zygotic hunchback mRNA bcd protein is transcribed at high levels at the anterior hb mRNA Review Bicoid protein regulates translation of caudal mRNA when bicoid protein is bound to the 3 UTR of caudal mRNA translational initiation factor cannot bind the 5 end of the caudal mRNA Gilbert 10e Fig 2 30 Caudal protein is present at higher levels in the posterior Gilbert 10e Fig 6 23 Nos protein also adopts a gradient mRNA nanos mRNA is tightly localized at the posterior nanos protein adopts a gradient protein courtesy Robin Wharton nanos is localized via oskar and regulates maternal hunchback translation nanos regulates maternal hb translation Gilbert 10e Fig 6 24 nanos regulates hunchback translation via pumilio ANTERIOR POSTERIOR Gilbert 8e Fig 9 21 Anterior pumilio protein bound to the 3 UTR of hunchback mRNA but no nanos translation of hunchback Posterior pumilio protein bound to the 3 UTR of hunchback mRNA nanos leads to deadenylation of hb mRNA and no hb protein Gap genes control a major region along the A P axis Kr ppel Gap genes are zygotically expressed in a wide band along the A P axis Mutants lack a large portion of the A P axis They all encode transcription factors Gilbert 10e Fig 6 27 Gap genes mutually repress one another Gilbert 9e Fig 6 28 Gap genes can mutually repress one another one gap protein causes loss of transcription of the mRNA encoding other gap proteins Posterior hunchback expression depends on terminal gap genes tailless and huckebein hunchback adapted from Gilbert 7e Fig 9 22 tailless tailless shown and huckebein not shown are terminal gap genes expressed at both ends of the zygote necessary for posterior hunchback expression Gap proteins have very specific zones of overlap hunchback Gilbert 10e Fig 6 18 Kr ppel Pair rule genes are expressed in 7 stripes fushi tarazu Pair rule genes are zygotically expressed in a 7 stripe pattern flies have 14 segments Their loss results in loss of every other segments or portions thereof They all encode transcription factors Gilbert 10e Fig 6 27 Pair rule gene example fushi tarazu Gilbert 10e Fig 6 33 Gap genes regulate pair rule genes Even skipped stripes correlate with zone of overlap of gap proteins hunchback hb Kr ppel Kr even skipped eve From FlyEx Even skipped stripes Reporter constructs Gilbert 10e Fig 6 31 Even skipped stripes are controlled by specific enhancer elements Reporter constructs show that specific enhancer elements control specific stripes Adapted from Gilbert 10e Fig 6 31 Bicoid gap genes regulate pair rule genes even skipped stripes Transcriptional activators bind to some enhancer elements activating transcription transcriptional repressors bind to some enhancer elements reducing transcription Regulators include bicoid and gap proteins Gilbert 7e Fig 9 24 Overlap of pair rule stripes Gilbert 10e Fig 6 30 even skipped fushi tarazu Pair rule proteins overlap with overlap as small as a few nuclei The complex patterns of overlap of pair rule proteins along with gap proteins results in 14 stripe patterns of expression segment polarity genes Overlap of pair rule stripes paired even