scholarly journals Adenomatous polyposis coli protein nucleates actin assembly and synergizes with the formin mDia1

2010 ◽  
Vol 189 (7) ◽  
pp. 1087-1096 ◽  
Author(s):  
Kyoko Okada ◽  
Francesca Bartolini ◽  
Alexandra M. Deaconescu ◽  
James B. Moseley ◽  
Zvonimir Dogic ◽  
...  
Keyword(s):  
Adenomatous Polyposis Coli ◽  
Microtubule Dynamics ◽  
Actin Dynamics ◽  
Adenomatous Polyposis ◽  
Actin Assembly ◽  
Polyposis Coli ◽  
Adenomatous Polyposis Coli Protein ◽  
Cell Protrusion

The tumor suppressor protein adenomatous polyposis coli (APC) regulates cell protrusion and cell migration, processes that require the coordinated regulation of actin and microtubule dynamics. APC localizes in vivo to microtubule plus ends and actin-rich cortical protrusions, and has well-documented direct effects on microtubule dynamics. However, its potential effects on actin dynamics have remained elusive. Here, we show that the C-terminal “basic” domain of APC (APC-B) potently nucleates the formation of actin filaments in vitro and stimulates actin assembly in cells. Nucleation is achieved by a mechanism involving APC-B dimerization and recruitment of multiple actin monomers. Further, APC-B nucleation activity is synergistic with its in vivo binding partner, the formin mDia1. Together, APC-B and mDia1 overcome a dual cellular barrier to actin assembly imposed by profilin and capping protein. These observations define a new function for APC and support an emerging view of collaboration between distinct actin assembly–promoting factors with complementary activities.

scholarly journals Adenomatous polyposis coli nucleates actin assembly to drive cell migration and microtubule-induced focal adhesion turnover

2017 ◽  
Vol 216 (9) ◽  
pp. 2859-2875 ◽  
Author(s):  
M. Angeles Juanes ◽  
Habib Bouguenina ◽  
Julian A. Eskin ◽  
Richa Jaiswal ◽  
Ali Badache ◽  
...  
Keyword(s):  
Cell Migration ◽  
Adenomatous Polyposis Coli ◽  
Focal Adhesions ◽  
Adenomatous Polyposis ◽  
Actin Assembly ◽  
Polyposis Coli ◽  
Directed Cell Migration ◽  
Direct Cell ◽  
Mitochondrial Distribution

Cell motility depends on tight coordination between the microtubule (MT) and actin cytoskeletons, but the mechanisms underlying this MT–actin cross talk have remained poorly understood. Here, we show that the tumor suppressor protein adenomatous polyposis coli (APC), which is a known MT-associated protein, directly nucleates actin assembly to promote directed cell migration. By changing only two residues in APC, we generated a separation-of-function mutant, APC (m4), that abolishes actin nucleation activity without affecting MT interactions. Expression of full-length APC carrying the m4 mutation (APC (m4)) rescued cellular defects in MT organization, MT dynamics, and mitochondrial distribution caused by depletion of endogenous APC but failed to restore cell migration. Wild-type APC and APC (m4) localized to focal adhesions (FAs), and APC (m4) was defective in promoting actin assembly at FAs to facilitate MT-induced FA turnover. These results provide the first direct evidence for APC-mediated actin assembly in vivo and establish a role for APC in coordinating MTs and actin at FAs to direct cell migration.

scholarly journals Actin/microtubule crosstalk during platelet biogenesis in mice is critically regulated by Twinfilin1 and Cofilin1

2020 ◽  
Vol 4 (10) ◽  
pp. 2124-2134 ◽  
Author(s):  
Isabelle C. Becker ◽  
Inga Scheller ◽  
Lou M. Wackerbarth ◽  
Sarah Beck ◽  
Tobias Heib ◽  
...  
Keyword(s):  
Regulatory Proteins ◽  
Actin Dynamics ◽  
Adenomatous Polyposis ◽  
Polyposis Coli ◽  
And Function ◽  
Redundant Functions ◽  
H Protein ◽  
Functional Defects

Abstract Rearrangements of the microtubule (MT) and actin cytoskeleton are pivotal for platelet biogenesis. Hence, defects in actin- or MT-regulatory proteins are associated with platelet disorders in humans and mice. Previous studies in mice revealed that loss of the actin-depolymerizing factor homology (ADF-H) protein Cofilin1 (Cof1) in megakaryocytes (MKs) results in a moderate macrothrombocytopenia but normal MK numbers, whereas deficiency in another ADF-H protein, Twinfilin1 (Twf1), does not affect platelet production or function. However, recent studies in yeast have indicated a critical synergism between Twf1 and Cof1 in the regulation of actin dynamics. We therefore investigated platelet biogenesis and function in mice lacking both Twf1 and Cof1 in the MK lineage. In contrast to single deficiency in either protein, Twf1/Cof1 double deficiency (DKO) resulted in a severe macrothrombocytopenia and dramatically increased MK numbers in bone marrow and spleen. DKO MKs exhibited defective proplatelet formation in vitro and in vivo as well as impaired spreading and altered assembly of podosome-like structures on collagen and fibrinogen in vitro. These defects were associated with aberrant F-actin accumulation and, remarkably, the formation of hyperstable MT, which appears to be caused by dysregulation of the actin- and MT-binding proteins mDia1 and adenomatous polyposis coli. Surprisingly, the mild functional defects described for Cof1-deficient platelets were only slightly aggravated in DKO platelets suggesting that both proteins are largely dispensable for platelet function in the peripheral blood. In summary, these findings reveal critical redundant functions of Cof1 and Twf1 in ensuring balanced actin/microtubule crosstalk during thrombopoiesis in mice and possibly humans.

scholarly journals Adenomatous polyposis coli protein (APC)-independent regulation of β-catenin/Tcf-4 mediated transcription in intestinal cells

1999 ◽  
Vol 344 (2) ◽  
pp. 565-570 ◽  
Author(s):  
Josep BAULIDA ◽  
Eduard BATLLE ◽  
Antonio GARCÍA DE HERREROS
Keyword(s):  
Adenomatous Polyposis Coli ◽  
Transcriptional Activity ◽  
Intestinal Cell ◽  
Adenomatous Polyposis ◽  
Polyposis Coli ◽  
Binding Assays ◽  
Adenomatous Polyposis Coli Protein ◽  
Kinase C ◽  
Intestinal Cell Lines

Alterations in the transcriptional activity of the β-catenin-Tcf complex have been associated with the earlier stages of colonic transformation. We show here that the activation of protein kinase C by the phorbol ester PMA in several intestinal cell lines increases the levels of β-catenin detected in the nucleus and augments the transcriptional activity mediated by β-catenin. The response to PMA was not related to modifications in the cytosolic levels of β-catenin and was observed not only in cells with wild-type adenomatous polyposis coli protein (APC) but also in APC-deficient cells. Binding assays in vitro revealed that PMA facilitates the interaction of the β-catenin with the nuclear structure. Our results therefore show that β-catenin-mediated transcription can be regulated independently of the presence of APC.

scholarly journals Interaction between Tumor Suppressor Adenomatous Polyposis Coli and Topoisomerase IIα: Implication for the G2/M Transition

2008 ◽  
Vol 19 (10) ◽  
pp. 4076-4085 ◽  
Author(s):  
Yang Wang ◽  
Yoshiaki Azuma ◽  
David Moore ◽  
Neil Osheroff ◽  
Kristi L. Neufeld
Keyword(s):  
Cell Cycle ◽  
Tumor Suppressor ◽  
Adenomatous Polyposis Coli ◽  
Fluorescent Protein ◽  
Adenomatous Polyposis ◽  
Polyposis Coli ◽  
Topoisomerase Iiα ◽  
Nuclear Localization Signals

The tumor suppressor adenomatous polyposis coli (APC) is implicated in regulating multiple stages of the cell cycle. APC participation in G1/S is attributed to its recognized role in Wnt signaling. APC function in the G2/M transition is less well established. To identify novel protein partners of APC that regulate the G2/M transition, APC was immunoprecipitated from colon cell lysates and associated proteins were analyzed by matrix-assisted laser desorption ionization/time of flight (MALDI-TOF). Topoisomerase IIα (topo IIα) was identified as a potential binding partner of APC. Topo IIα is a critical regulator of G2/M transition. Evidence supporting an interaction between endogenous APC and topo IIα was obtained by coimmunoprecipitation, colocalization, and Förster resonance energy transfer (FRET). The 15-amino acid repeat region of APC (M2-APC) interacted with topo IIα when expressed as a green fluorescent protein (GFP)-fusion protein in vivo. Although lacking defined nuclear localization signals (NLS) M2-APC predominantly localized to the nucleus. Furthermore, cells expressing M2-APC displayed condensed or fragmented nuclei, and they were arrested in the G2 phase of the cell cycle. Although M2-APC contains a β-catenin binding domain, biochemical studies failed to implicate β-catenin in the observed phenotype. Finally, purified recombinant M2-APC enhanced topo IIα activity in vitro. Together, these data support a novel role for APC in the G2/M transition, potentially through association with topo IIα.

Μελέτη των μηχανισμών εκδήλωσης συγγενών καρδιοπαθειών με την χρήση του zebrafish ως πειραματικό μοντέλο

2016 ◽  
Author(s):  
Δέσποινα Μπουρνελέ
Keyword(s):  
Adenomatous Polyposis Coli ◽  
Outflow Tract ◽  
Adenomatous Polyposis ◽  
Polyposis Coli

Οι βαλβιδοπάθειες με συχνότητα εμφάνισης 5% επί των ζώντων νεογνών αποτελούν το 20-30% του συνόλου των συγγενών καρδιοπαθειών. Οι καρδιακές βαλβίδες αναπτύσσονται και διαμορφώνονται ταυτόχρονα με το σχηματισμό της καρδιάς και όσο αυτή συστέλλεται. Οι βαλβίδες λειτουργούν καθ’όλη τη διάρκεια της ζωής ενός οργανισμού με σκοπό την παρεμπόδιση της παλίνδρομης ροής του αίματος μεταξύ των καρδιακών κοιλοτήτων. Η ακριβής τους διαμόρφωση είναι σημαντική για την φυσιολογική καρδιακή λειτουργία και είναι αποτέλεσμα της αλληλεπίδρασης της συσταλτικότητας της καρδιάς και της αιμοδυναμικής ροής. Ποικίλα σηματοδοτικά μονοπάτια εμπλέκονται στη μορφοποίηση των καρδιακών βαλβίδων. Ωστόσο, οι μοριακοί μηχανισμοί που διέπουν την ανάπτυξη τους δεν έχουν πλήρως διαλευκανθεί.Το zebrafish αποτελεί ένα ιδανικό πειραματικό μοντέλο για τη μελέτη της ανάπτυξης των καρδιακών βαλβίδων καθώς επιτρέπει τη μη επεμβατική in vivo παρατήρηση της ανάπτυξης του καρδιαγγειακού συστήματος. Επίσης, η καρδιά του, zebrafish έχει την ικανότητα να αναγεννάται σε όλη τη διάρκεια ζωής του ψαριού, προσφέροντας γνώση για την κατανόηση των μηχανισμών που διέπουν την αναγέννηση της ανθρώπινης καρδιάς. Το zebrafish είναι ακόμα ένα πολύτιμο γενετικό εργαλείο για την ταυτοποίηση υποψήφιων γονιδίων για συγγενείς καρδιοπάθειες, είτε μέσω του χαρακτηρισμού μεταλλαγμένων σειρών που έχουν προέλθει από γενετικούς ελέγχους είτε μέσω της φαινοτυπικής παρατήρησης εμβρύων μετά από εφαρμογή μεθόδων γονιδιωματικής τροποποίησης.Στην παρούσα μελέτη πραγματοποιήθηκε η ταυτοποίηση και ο χαρακτηρισμός γονιδίων που εμπλέκονται στην ανάπτυξη και το σχηματισμό των καρδιακών βαλβίδων. Οι μεταλλαγμένες γενετικές σειρές που χρησιμοποιήθηκαν προήλθαν από τυχαία μεταλλαξιγένεση στα πλαίσια ενός ελέγχου πρόσθιας γενετικής για καρδιοαγγειακούς φαινότυπους. Η μεταλλαγμένη σειρά bua εμφανίζει στένωση της βαλβίδας στο σημείο εξώθησης της καρδιάς (outflow tract), με αποτέλεσμα την ελαττωματική ανάπτυξη της κολποκοιλιακής βαλβίδας και την παλινδρόμησή του αίματος μεταξύ κόλπου και κοιλίας. Τα έμβρυα της μεταλλαγμένης σειράς BH εμφανίζουν μικρότερη σε μέγεθος καρδιά και κυρτή ουρά. Παράλληλα, πραγματοποιήθηκε ο χαρακτηρισμός της διαγονιδιακής σειράς Tg(7xTCF-Xla.Siam:nlsmCherry). Πρόκειται για μια νέα διαγονιδιακή σειρά αναφοράς της Wnt/β-κατενίνης, που συμβάλλει στον εντοπισμό της ενεργότητας του σηματοδοτικού μονοπατιού στα κύτταρα του zebrafish. Ο χαρακτηρισμός του διαγονιδίου πραγματοποιήθηκε και στη μεταλλαγμένη σειρά apchu745, στην οποία το σηματοδοτικό μονοπάτι Wnt/β-catenin είναι μόνιμα ενεργοποιημένο. Έμβρυα zebrafish με μεταλλάξεις στο ογκοκατασταλτικό γονίδιο apc (adenomatous polyposis coli) εμφανίζουν ανωμαλίες στην ανάπτυξη της καρδιάς, όπως ελαττωματική καρδιακή συσταλτικότητα, ελαττωματική κάμψη της καρδιάς, καθώς επίσης και υπερπλαστικά καρδιακά επάρματα στις καρδιακές βαλβίδες.Σκοπός της συγκεκριμένης εργασίας ήταν η μελέτη της λειτουργίας των γονιδίων, η ταυτοποίηση νέων σηματοδοτικών μονοπατιών και η διαλεύκανση των μοριακών μηχανισμών που εμπλέκονται στην παθογένεση των συγγενών καρδιοπαθειών. Αρκετά μοναδικά χαρακτηριστικά καθιστούν το zebrafish ένα ελκυστικό πειραματικό μοντέλο, που προσφέρει τη δυνατότητα μοντελοποίησης μεγάλου εύρους ανθρώπινων ασθενειών.

scholarly journals The Adenomatous Polyposis Coli Protein Is Required for the Formation of Robust Spindles Formed in CSF Xenopus Extracts

2004 ◽  
Vol 15 (6) ◽  
pp. 2978-2991 ◽  
Author(s):  
Dina Dikovskaya ◽  
Ian P. Newton ◽  
Inke S. Näthke
Keyword(s):  
Adenomatous Polyposis Coli ◽  
Chromosomal Instability ◽  
Adenomatous Polyposis ◽  
Mitotic Spindles ◽  
Polyposis Coli ◽  
Adenomatous Polyposis Coli Protein ◽  
Microtubule Binding ◽  
Microtubule Polymerization ◽  
Large N ◽  
Apc Protein

Mutations in the adenomatous polyposis coli (APC) protein occur early in colon cancer and correlate with chromosomal instability. Here, we show that depletion of APC from cystostatic factor (CSF) Xenopus extracts leads to a decrease in microtubule density and changes in tubulin distribution in spindles and asters formed in such extracts. Addition of full-length APC protein or a large, N-terminally truncated APC fragment to APC-depleted extracts restored normal spindle morphology and the intact microtubule-binding site of APC was necessary for this rescue. These data indicate that the APC protein plays a role in the formation of spindles that is dependent on its effect on microtubules. Spindles formed in cycled extracts were not sensitive to APC depletion. In CSF extracts, spindles predominantly formed from aster-like intermediates, whereas in cycled extracts chromatin was the major site of initial microtubule polymerization. These data suggest that APC is important for centrosomally driven spindle formation, which was confirmed by our finding that APC depletion reduced the size of asters nucleated from isolated centrosomes. We propose that lack of microtubule binding in cancer-associated mutations of APC may contribute to defects in the assembly of mitotic spindles and lead to missegregation of chromosomes.

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